Hydrosphere

To better understand the adsorption of each drug in the different samples, a kinetic assay was performed for 48 h. The methodology consisted of weighing 2.0 ± 0.1 g of each CNT sample, in Eppendorf tubes and putting them in contact with a solution of each drug, individually, at 250 mg∙dm⁻³ with methanol (CH_3OH, VWR CHEMICALS, ≥99.9%) for [0:00; 0:15; 0:30; 1:00; 2:00; 4:00; 18:00; 24:00; 47:00] h. The Eppendorf tubes were put on a P SELECTA ROTABIT shaker at 130 rpm. Each Eppendorf tube was removed and placed in a VWR MicroStar12 centrifuge at 30 rpm for 15 min. Finally, ≈0.8 cm⁻³ of the supernatant was removed and placed in vials, which were then read using an analytical method, high-performance liquid chromatography (HPLC). HPLC methods were optimized for each drug based on the literature [36 ,37 (link),38 (link)] and performed in equipment (Hitachi Elite LaChrom HPLC, Tokyo, Japan) equipped with a UV detector and a C₁₈ column (YMC Hydrosphere, 250 mm × 4.6 mm). Thus, the method selected involved a ratio of 20% sodium phosphate buffer solution monobasic monohydrate at 0.05 M (H₂NaO₄P-H₂O, SIGMA-ALDRICH, ≥98%) and 80% acetonitrile (CH₃CN, VWR CHEMICALS, ≥99.95%); an injection volume of 10 µL in TAC and 20 µL in ETA and 5-FU; at 1 mL∙min⁻¹; and wavelengths for TAC, ETA, and 5-FU of 243, 254, and 266 nm, respectively.

All peptides were synthesized as light and heavy pairs by 21st Century Biochemicals (Marlborough, MA). Heavy peptides were labeled with C-terminal R [¹³C₆¹⁵N₄] or K [¹³C₆¹⁵N₂] with > 99% isotopic enrichment. For cysteines, carbamidomethylated cysteines were used. Synthesized peptides were further purified by reverse-phase HPLC, using a YMC Hydrosphere 20 × 150 mm C18 column, packed with 5 micron beads with 120 Å pore size, to attain > 95% purity, and net peptide content was verified by amino acid analysis.