Hydrochloric acid (hcl)

2.0 × 10⁴ (4 h), 1.5 × 10⁴ (24 h), 1.0 ×x 10⁴ (48 h) and 6.5 × 10³ (72 h) cells were treated with CAP or carrier gas argon by kINPen (15 s) or Mini Jet (30 s) and were incubated for 4, 24, 48, and 72 h. To be able to normalize the metabolized MTT to the cell number, a second plate was planted out parallel. For performing the MTT-Assay, 20 µL MTT-Solution (MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Carl Roth, Karlsruhe, Germany) in a. bidest (5mg/mL) was added and incubated over 3 h. After incubation, the culture media were removed, and 120 µL lysis reagent (DMSO (Carl Roth, Karlsruhe, Germany) with 2.5% w/w SDS (Sodium Dodecyl Sulfate) (Carl Roth, Karlsruhe, Germany) and 150 µL 37% HCl (Carl Roth, Karlsruhe, Germany) were added and incubated over 10 min. Absorption at 565 nm was measured using an Infinite M200 plate reader (Tecan, Männedorf, Switzerland).

Sugar beet pectin was provided by Herbstreith & Fox KG. (Neuenbürg, Germany). The main molecular characteristics are protein content: 4.1% ± 0.2%; molecular weight: 104 kDa ± 5 kDa; galacturonic acid: 66.1% ± 1.8%; degree of methylation: 52.1% ± 1.7%; degree of acetylation: 23.1% ± 0.5%; and trans-ferulic acid: 706 mg/100g ± 21 mg/100 g [18 (link)].
Medium chain triglyceride oil (MCT oil) was supplied by IOI Oleo GmbH (Hamburg, Germany). The MCT oil was composed of 60% C₈ and 40% C₁₀ chains and had a density of 0.95 kg/L at room temperature, according to supplier information. 1-Dodecanthiol (≥98%), hydrogen peroxide (30%), and sulfuric acid (98%) were purchased from Merck KGaA (Darmstadt, Germany). Sodium chloride, hydrochloric acid, sodium hydroxide, and Florisil were obtained from Carl Roth GmbH & Co. KG. (Karlsruhe, Germany). All chemicals were at least of analytical grade, and ultrapure water was used throughout the experiments.

Acetonitrile, chloroform, methanol, acetic acid, phosphoric acid, and hydrochloric acid were purchased from Carl Roth GmbH & Co KG (Karlsruhe, Germany). Ammonium acetate, copper sulfate, and potassium chloride were purchased from Sigma-Aldrich GmbH (Munich, Germany). The standard substances SQDG 816 (2-O-hexadecanoyl-1-O-(9Z,12Z,15Z-octadecatrienoyl)glycerol 3-(6-deoxy-6-sulfo-α-d-glucopyranoside)) and 3-O-sulfo-d-galactosyl-β1-1′-N-heptadecanoyl-d-erythro-sphingosine as internal standard (ISD) were purchased from Avanti Polar Lipids Inc. (Alabaster, AL, USA). All aqueous solutions were prepared with deionized water, generated by a Purelab flex water purification system (Veolia Water Technologies Deutschland GmbH, Celle, Germany). NH₂ cartridges (6 mL, 500 mg) were purchased from Macherey-Nagel GmbH & Co. KG (Düren, Germany).

1,4-piperazinediethanesulfonic acid (PIPES), histamine dihydrochloride, sodium hydroxide (NaOH), D(+)-sucrose, monobasic potassium phosphate (KH₂PO₄), hydrochloric acid (HCl), and hydrogen peroxide (30%) were purchased from Carl Roth GmbH (Karlsruhe, Germany). Sodium dihydrogen phosphate, sodium diethyldithiocarbamate, ortho-phosphoric acid (H₃PO₄), and thiamine chloride dihydrochloride were purchased from Merck (Darmstadt, Germany). Bovine serum albumin (BSA; modified Cohn Fraction V, pH 5.2) was purchased from Serva electrophoresis GmbH (Heidelberg, Germany). Catalase (from Micrococcus lysodeikticus; 111,700 U·mL⁻¹) and pancreatin from porcine pancreas (8× USP specifications) were purchased from Sigma-Aldrich (Merck) (St. Louis, MO, USA). (10-(carboxymethyl-aminocarbonyl)-3,7-bis(dimethylamino) phenothiazine sodium salt (DA-67) was purchased from Fujifilm Wako Chemicals U.S.A. Corp (Richmond, VA, USA). Horseradish peroxidase (Grade I) was purchased from AppliChem GmbH (Darmstadt, Germany). Whey protein isolate (WPI; 90% (w/w) protein) was obtained from Sachsenmilch Leppersdorf GmbH (Leppersdorf, Germany). Sodium caseinate (90.6% (w/w) protein) was obtained from FrieslandCampina (Amersfoort, Netherlands).

Pure standard of caffeine, myristic acid (99%), palmitic acid (99%), acetyl chloride (98%), trolox (97%), and ethanol (96 vol%, not denaturated) were obtained from ACROS ORGANICS. Acetonitrile, ethyl acetate, and potassium hydroxide were supplied by Fisher Chemical (Waltham, MA, USA). Pure standard of 3-caffeoylquinic acid (CAS 327-97-9), oleic acid (99%), stearic acid (98%), Vitamin E acetate (97%), DPPH (95%), and cyclohexane were purchased from Alfa Aesar (Haverhill, MA, USA). Caprylic acid (99%) and arachidic acid (99%) were obtained from Sigma Aldrich (St. Louis, MO, USA). Capric acid (99%) and lauric acid (99%) were obtained from Interchim (Montlucon, France). Methanol (≥99.9%) was obtained from Honeywell (Nawabash, IN, USA). Hydrochloric acid (37%) was obtained from Roth. Linoleic acid (99%), and n-hexane (>99.9%) were obtained from Fluka. All solvents and reagents were of analytical grade and used as received.

For determination of recombinant NOS2 enzyme activity, 2 units of recombinant enzyme (Biomol, Hamburg, Germany) were incubated in appropriate buffer following the manufacturer’s protocol. In short, 50 mM HEPES (Carl Roth, Karlsruhe, Germany) and 1 mM magnesium acetate (Sigma Aldrich) were adjusted to pH 7.4 or 6.0 by addition of hydrochloric acid (Carl Roth). Prior to buffer use, 0.15 mM NADPH (Sigma Aldrich), 4.5 µM oxyhemoglobin (Sigma Aldrich), 18 µM tetrahydrobiopterin (Sigma Aldrich), and 180 µM dithiothreitol (Sigma Aldrich) were freshly added. 1 mM Arg-HCl (Sigma Aldrich) was also freshly added where indicated. After incubation at 37°C, nitrite accumulation was quantified at distinct time points between 1 h and 8.5 h using the Griess assay (32 (link)).