Sodium carbonate anhydrous

Pectinase from Aspergillus aculeatus (Pectinex® Ultra Clear, a blend of pectinases, hemicellulases and arabinases with activity of 8600 PGNU/g) and (Pectinex® Ultra SP-L, a blend of pectinases, hemicellulases and beta-glucanases with activity of 3300 PGNU/g), cellulase from Trichoderma reesei (Celluclast® 1.5L, a cellulase aqueous solution with activity of ≥700 units/g) were obtained from Sigma Aldrich (Darmstadt, Germany). Information about the optimal conditions of these enzymes as declared by the producer can be found in Table 1. Microcrystalline cellulose Avicel® PH-101, pectin from citrus peel, 3,5-dinitro salicylic acid, D-(+)-galacturonic acid monohydrate and D-(+)-glucose were purchased from Sigma Aldrich (Darmstadt, Germany). Sodium carbonate anhydrous, citric acid, sodium hydroxide, and L (+)-potassium sodium tartrate tetrahydrate were purchased from VWR Chemicals (Leuven, Belgium), while di-sodium hydrogen phosphate anhydrous, sodium acetate trihydrate, and acetic acid 99–100 % were obtained from Chem-Lab (Zedelgem, Belgium).Table 1

Conditions range of activity of the used enzymes as declared by the producer.

Enzyme	pH range of activity	Temperature range of activity
Pectinex® Ultra Clear	1.8–3.0	25–40 °C
Pectinex® Ultra SP-L	2.8–6.5	15–55 °C
Celluclast® 1.5L	4.0–6.0	50–60 °C

Formic acid (LC–MS grade) was obtained from Merck KGaA (Darmstadt, Germany). Acetonitrile and mEthanol (both Optima® LC-MS grade) were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Extracts were filtered through a 0.20 μm, 4 mm titan syringe filter (Thermo Fisher Scientific). Ultra-pure water was obtained from a water purification system (Millipore Direct-Q3-UV, Merck KGaA, Darmstadt, Germany). Ethanol, hexane, glacial acetic acid and sodium carbonate anhydrous were obtained from Avantor Performance Materials Poland (Gliwice, Poland). Gallic, oleanolic acid, β-carotene, vanillin, 2,2′-azino-bis-3(ethylbenzthiazoline-6-sulphonic acid) (ABTS) and Trolox were purchased from Sigma-Aldrich Chemical Co. (St. Louis, Mo, USA). Perchloric acid and Folin-Ciocalteu reagent were obtained from Chempur (Piekary Śląskie, Poland).

Ethanol, Folin–Ciocalteu reagent, (Merck, Darmstadt, Germany), 3,5-dinitrosalicylic acid, 2,2-diphenyl-1-picrylhydrazyl (DPPH), 4-nitrophenyl α-D-glucopyranoside (PNPG), α-glucosidase, α-amylase, acetonitrile, collagenase from Clostridium histolyticum, formic acid, iron (II) chloride tetrahydrate, ferrozine, N-[3-(2-Furyl)acryloyl]-Leu-Gly-Pro-Ala (FALGPA), neocuproine, tricine, (Sigma-Aldrich, St. Louis, MO USA); aluminum chloride anhydrous, ammonium acetate, calcium chloride, copper (II) chloride dihydrate, disodium hydrogen phosphate dodecahydrate, methanol, sodium carbonate anhydrous, sodium dihydrogen phosphate dehydrate, sodium chloride, sodium hydroxide, and sodium phosphate monobasic (Avantor Performance Materials Poland S.A., Gliwice, Poland), were used. Standards: gallic acid and acarbose, ethylenediaminetetraacetic acid (EDTA) (Sigma-Aldrich, St. Louis, MO USA), chlorogenic acid, protocatechuic acid, quercetin, and rutin (Carl Roth GmbH + Co. KG, Karlsruhe, Germany) were used.

Poly(ethylene glycol) (PEG; MW 10,000, 600, 400 and 200 Da), methylene blue and HPLC grade methanol and acetonitrile were purchased from Sigma–Aldrich (Dorset, UK). ATR, NR, OLP and propylene glycol were purchased from Tokyo Chemical Industry UK Ltd. (Oxford, UK). Gantrez^® S-97, a copolymer of methyl vinyl ether and maleic acid (PMVE/MA, molecular mass: 1,500,000 Da) was a gift from Ashland (Kidderminster, UK). Anhydrous sodium carbonate was obtained from VWR International Ltd., (Poole, UK). Oxoid™ phosphate buffered saline (PBS) pH 7.4 tablets were obtained from Thermo Scientific (Sheffield, UK). Poly(ester) film, one-side siliconised, release liner, FL2000 PET 75 was purchased from Rexam Release B.V (Apeldoom, The Netherlands) Glisseal^®N vacuum grease was purchased from Borer Chemie (Zuchwil, Switzerland). All other chemicals used were of analytical reagent grade.

Anhydrous sodium carbonate (>99.5%) was purchased from VWR International s.r.l. (Milan, Italy, while the nitrite and nitrate certified anion standard solutions for ion chromatography (1000 mg L⁻¹) were supplied by Sigma-Aldrich (Stenheim, Germany). The solvent used for standards and mobile phase preparation and sample extraction was ultrapure water, with a specific resistance of 18.2 MΩ cm⁻¹, supplied by an Arium mini essential UV system (Sartorius AG, Goettingen, Germany). The sodium carbonate solution, used as the mobile phase, was filtered before use through 0.45 μm membrane filters and degassed with helium. The stock solutions of nitrite and nitrate at a concentration of 1000 mg L⁻¹ were opportunely diluted for calibration purposes of the traditional ion chromatography method (HPIC) to provide nitrite concentrations of 0.2, 0.4, 0.8, 1.6, and 6.5 mg L⁻¹ and nitrate concentrations of 0.6, 1.2, 2.4, 4.8, and 12.5 mg L⁻¹. The calibration levels used for the confirmatory technique, capillary ion chromatography (CIC), were 0.02, 0.1, 0.2, 2.0, and 20 mg L⁻¹ for both nitrite and nitrate.