Alginic acid sodium salt from

Alginate solution is prepared freshly for the next day. Sodium alginate powder (Vivapharm alginate PH176, batch nr. 4503283839, JRS Pharma GmbH, Rosenberg, Germany, or alginic acid sodium salt from brown algae, A0682, Sigma Aldrich, for THP1 cells) is dispersed at a concentration of 1.5%, 2%, or 2.5% (w/v) in serum-free cell culture medium (Table 1). The alginate solution is mixed overnight with a magnetic stirrer at room temperature until all powder has been dissolved. The suspension fluid is then equilibrated by incubating for 6 hr at 37 °C, 5% CO₂. When prepared with RPMI media (but not when prepared with DMEM nor Sigma Aldrich alginate), the alginate solution is filtered with a 0.45 µm filter before use. 1 ml of alginate solution are then added to the cell pellet of 10⁶ cells in the Falcon tube and mixed using a positive displacement pipette (15314274, Gilson/Fisher Scientific) by slowly (~2 s cycle time) and repeatedly (10 x) sucking the liquid in and out. The alginate-cell suspension is then transferred into a 2 ml screw-cup test tube and centrifuged for 30 s at 150 rcf to remove air bubbles.

Alginic acid sodium salt from brown algae (SA) (medium viscosity), HA with a molecular formula of Ca₅(OH)(PO₄)₃, and calcium chloride (CaCl₂) (pure reagent > 99%) were purchased from Sigma-Aldrich. For the elaboration of the hydrogels, different concentrations of HA NPs were used. The samples were named “SA(NPs)-(%NPs)” where (NPs) means the NPs used and (%NPs) represents the percentage of the NPs in the total weight between SA and NPs. The different samples prepared for this study are shown in Table 1.
The selected concentration of NPs was dispersed in distilled water (the amount was determined by the mass of the SA, 25 mL for every gram of SA) by magnetic stirring over 25 min. For better dispersion of the NPs, ultrasonication was applied for 5 min with a Cole-Parmer 500-Watt ultrasonic homogenizer with a frequency of 20 kHz. Then, the corresponding amount of SA was added to the blend and left in magnetic stirring overnight. Once the blend showed a homogeneous mixture, the hydrogel was cross-linked with 5 wt.% CaCl₂ in a 0.5 M solution, where hydrogels went from liquid to solid state.

Met HCl was purchased from Farmhispania, S.A. (Barcelona, Spain). Alginic acid sodium salt from brown algae with medium viscosity (Sod ALG), sodium chloride, and hydrochloric acid fuming 37% were obtained from Sigma-Aldrich (St. Louis, MO, USA). Natrosol™ 250 hydroxyethylcellulose (HEC) HHX grade was purchased from Ashland (Covington, KY, USA). Calcium carbonate and sodium carbonate (USP grade) were obtained from Daejung (Siheung, Korea). Gaviscon Double Action was purchased from Oxy Reckitt Benckiser (Seoul, Korea). Purified water was prepared using a Milli-Q^® Direct 8 Water Purification System from EMD Millipore (Darmstadt, Germany). HPLC grade solvents were used for high-performance liquid chromatography.