Milli q ultrapure water

All thiolated oligonucleotides used in this study were purchased from Integrated DNA Technologies (Coralville, IA). Graphite powder (99%, 7–11 μM) was purchased from Alfa–Aesar (Heysham, Lancashire, UK). Sulfuric acid, phosphoric acid, hydrogen peroxide, trisodium citrate, sodium phosphate tribasic, sodium phosphate dibasic, boric acid, tetrachloroauric acid, dimethyl sulfoxide, apigenin, xylene, ethanol, 3,3-diaminobenzidine tetrahydrochloride, hematoxylin, and isopropanol were purchased from Sigma–Aldrich (Milwaukee, WI). Citric acid, calcium chloride, magnesium chloride, potassium permanganate, tris(hydroxymethyl)aminomethane (Tris), and hydrochloric acid were purchased from Mallinckrodt Baker (Phillipsburg, NJ). Alpha-MEM and fetal bovine serum were purchased from GIBCO (Campinas, Brazil). An MTT assay kit was purchased from Thermo Fisher Scientific (Waltham, MA). MUC1 (VU4H5) mouse mAb, anti-mouse IgG, and HRP-linked antibody were purchased from Cell Signaling Technology (Danvers, MA). Milli-Q ultrapure water (18 MΩ, Millipore, Billerica, MA) was used in all experiments.

Stock solutions of metal contaminants (Pb, Cd) were prepared at six different concentrations (10, 20, 40, 60, 80, and 100 mg L⁻¹) using Pb and Cd salts (PbCl₂, CdCl₂ × H₂O) and Milli-Q ultrapure water (Millipore, Bedford, MA, USA) with pH adjusted to 5.0. The effects of two parameters (initial metal concentration and contact time) were investigated to study the adsorptive behavior of CHIT-PAAA. Pb and Cd adsorption assays were performed on the synthesized material CHIT-PAAA under magnetic agitation (at 600 rpm rotational speed) and normal atmospheric conditions (room temperature). Afterwards, samples were filtered off (on Rotilabo folded filters, type 113 P, membrane Ø 150 mm, Macherey-Nagel GmbH, Dueren, Germany), and heavy metals in the supernatant were analyzed by FAAS. An AAS Spectra AA110 atomic absorption spectrophotometer was used to determine the metal concentrations in the solutions.
The removal efficiencies (adsorption percentages) and sorption capacities were calculated based on the following equations: $$\mathrm{R} (\%)=\frac{{\mathrm{C}}_{\mathrm{i}}-{\mathrm{C}}_{\mathrm{f}}}{{\mathrm{C}}_{\mathrm{i}}}· 100$$
$$\mathrm{q} \left({\mathrm{mg} \mathrm{g}}^{-1}\right)=\frac{\left({\mathrm{C}}_{\mathrm{i}}-{\mathrm{C}}_{\mathrm{f}}\right)· \mathrm{V}}{\mathrm{w}}$$
where R is the removal efficiency (%); C_i is the initial concentration (before adsorption) (mg L⁻¹); C_f is the final concentration (after adsorption) (mg L⁻¹); q is the sorption capacity (mg g⁻¹); V is the volume of solution (L); w is the amount of sorbent (material) used (g).

All chemicals are of analytical reagent grade and were used as-received without further purification. Citric acid, ammonia water, methanol, ethyl alcohol 3-(2-aminoethylamino)-propyldimethoxymethylsilane(AEAP),(3aminopropyl)triethoxysilane (APTES), tetraethyl orthosilicate (TEOS), naproxen, ibuprofen, ketoprofen, Fenoprofen were obtained from Saen Chemical Technology (Shanghai) Co., Ltd. Millipore Milli-Q ultrapure water was used for all the experiments in the work.

The samples (100-200 mg) were immediately frozen in liquid nitrogen, then freeze-dried, and their dry weight was determined. For extraction, the samples were boiled with 0.5 mL of 80% methanol (v/v) at 75 °C for 15 min, the solvent was vacuum-evaporated, and the residue was resuspended in Milli-Q ultrapure water (Millipore). The content of non-structural soluble carbohydrates was determined using high performance liquid chromatography (HPLC, flow rate 0.5 mL min⁻¹, 80 °C) with refractometric detection (refractive index range 1–1.75; refractometer Shodex RI-71; Spectra Physics—Newport Corporation, Irvine, CA, USA), column: IEX Ca²⁺ (Shodex). The starch in the pellets after the extraction of soluble carbohydrates was hydrolyzed by α-amylase (Fluka, 30U) and amyloglucosidase (Sigma, 60U) in 0.1 M Na-acetate buffer (pH 4.5), and the glucose content was measured by HPLC. The data were evaluated using Clarity 7.2 software (DataApex).

Carbamazepine (CBZ, >98% purity),
sulfadimethoxine (SDM, >98% purity), trimethoprim (TMP, >98%
purity),
all test amino acids (i.e., glycine, alanine, serine, leucine, proline,
phenylalanine, glutamic acid, asparagine, and histidine), phenyl methyl
sulfoxide (PMSO), phenyl methyl sulfone (PMSO₂), sodium
thiosulfate, and sodium borate were purchased from Fisher Scientific
(Austin, Texas, USA). A wet chemical procedure was applied to prepare
solid potassium ferrate(VI) (K₂FeO₄),^{59 (link)} which had a purity of more than 90%. The solid
K₂FeO₄ was dissolved in a 0.01 M sodium borate
buffer solution. The concentration of Fe(VI) solution was determined
by measuring the absorbance at 510 nm using a 1.0 cm path length and
a molar absorption coefficient of ε_510nm = 1150 M^–1 cm^–1,^{59 (link)} with a UV–visible spectrometer (Thermo Scientific Co., USA).
All reaction solutions were prepared using Milli-Q ultrapure water
(>18 MΩ cm^–1 resistivity, Millipore, Milford,
USA).

MilliQ ultrapure water, obtained by means of a Millipore equipment (Bedford, MA, USA) was used throughout. All chemicals employed for the extract preparations and for the subsequent analysis were HPLC-grade. They were supplied by VWR International S.r.l (Milan, Italy) and Sigma-Aldrich (St. Louis, MO, USA). Purity of all the used standards for HPLC analysis of BDs has been reported in the Supplementary Materials (Table S1).

E1, E2, EE2, and E3 (purity > 97%) were purchased from Sigma-Aldrich (Munich, Germany). Methanol and acetonitrile (HPLC grade) were obtained from Burdick & Jackson (Honeywell, Morristown, NJ, USA). NaOH (AR) and HCl (AR) were purchased from the Beijing Chemical Engineering Factory (Beijing, China). CaCl₂ was purchased from the Xilong Chemical Plant of Shantou, Guangdong. NaN₃ (AR) was obtained from the Tianjin Fuchen Chemical Reagents Factory. Sediments were collected from the Songhua River in the Jilin Province, China.
A 1200-HPLC (Agilent Company, Santa Clara, CA, USA), equipped with two model pumps (G1312A), an in-line degasser (G1322A), a column oven (G1316A), and a fluorescence detector (G1321A), was used for HPLC analysis. The injection loop volume was 20.0 µL, and a Zorbox SB-C18 column (250 mm × 4.6 mm; 5 µm) was used for the separations. A 2-16K centrifuge (Sigma, Munich, Germany), a FA-1004 analytical balance (Shanghai Hengping Science Instrument Company, Shanghai, China), and Milli-Q ultrapure water (Millipore, Billerica, MA, USA) were also used.