Agilent 240 aa

Due to possible emission spectrum interferences between Ag and Mg, element releases (Ag and Mg) during degradation were quantified either via AAS (for Mg samples) or ICP-MS (for Mg–6Ag samples). Prior to AAS analysis, the supernatants were acidified with 1 % (w/v) nitric acid (HNO₃, Merck KGaA, Darmstadt, Germany) and kept at 4 °C to avoid precipitation. Then, samples were diluted 1:250 in 1% HNO₃ in ultrapure H₂O, and total Mg concentration was detected using a flame AAS (Agilent 240 AA, Agilent Technologies, Waldbronn, Germany) at a wavelength of 285.2 nm (emission spectrum of Mg). A calibration curve ranging from 0.05 to 1 mg/L Mg was applied for quantification. ICP-MS was applied to quantify the concentration of Mg and Ag in the supernatant after Mg–6Ag degradation. To avoid contaminations, digiTUBEs (S-prep GmbH, Überlingen, Germany) were flushed twice with ultrapure water and 1% (w/v) HNO₃ in ultrapure water. Supernatants acidified in 1% (w/v) HNO₃ were diluted 1:1000–2000 in 1% (w/v) HNO₃ in ultrapure water to a final volume of 30 mL in digiTUBES. The concentrations of Mg and Ag were determined by an inductively-coupled plasma-mass spectrometer (Agilent 7900 ICP-MS, Agilent Technologies, Waldbronn, Germany) with an ESI PFA microflow nebulizer (Elemental Scientific, Omaha, NE, USA).

An in vitro immersion test calculating the weight loss after 72 h immersion under cell culture conditions (37 °C, 5% CO₂, 20% O₂, humidified atmosphere) was performed as described in Nidadavolu [53 (link)] in order to determine the impact of liposomes on the degradation of the Mg-10Gd material. Material discs 1.5 mm in height and 1 cm in diameter were cleaned utilizing 20 min sonication in n-hexane (Merck, Darmstadt, Germany), 20 min sonication in acetone (Merck, Darmstadt, Germany), and 3 min sonication in 100% ethanol. Lastly, specimens were sterilized and dried in 70% ethanol under sterile conditions [54 (link),55 (link)].
The discs were placed in Dulbecco’s modified eagle’s medium (DMEM, Life Technologies, Darmstadt, Germany) + 10% FBS (fetal bovine serum, PAA Laboratories, Linz, Austria): αMSH-SM-liposome (25:1 v:v), and in DMEM and DMEM 10% FCS: PBS (phosphate-buffered saline) (25:1 v:v). The Mg concentration released into the immersion medium was measured utilizing atomic absorption spectrometry (Agilent 240 AA, Agilent, CA, USA), as described [54 (link)]. The immersion assay was also performed with pure Mg using the same methodology.