5 hmf

Raw materials, including cellulose, D-allose (C₆H₁₂O₆), D-glucose (C₆H₁₂O₆), xylan, 5-HMF (C₆H₆O₃), LGA (C₆H₁₀O₅) and LGO (C₆H₆O₃), were reagent grade with a purity >99% and were purchased from Merck Co Ltd. The chemicals for catalyst synthesis, including Pd(NO₃)₂·2H₂O (~40% Pd basis), ZnSO₄·H₂O (≥99.9% trace metals basis) and ZnSO₄·7H₂O (ACS reagent, 99%), were also purchased from Merck Co Ltd. The two real biomass feedstocks, corncob and sugarcane bagasse with particle sizes of 150–250 μm, were sourced from Oz Gun Mart in New South Wales, Australia and Sugarcane Juice Bar in Melbourne, Australia, respectively. They were dried in an oven at 105 °C for 15 h prior to the pyrolysis experiments.

After surgery, animals were kept under normoxia for 30 min, after which baseline measurements were taken. Then, twelve animals (N = 12) were randomly assigned to receive either 100 uL of sterile saline with 100 mg/kg 5-HMF (Sigma-Aldrich, St. Louis, MO, United States) or only saline 20 min before exposure to hypoxia. This dosage is based in previous in vivo mice studies, where a single 100 mg/kg oral dose was efficacious in prolonging survival under severe hypoxia (Abdulmalik et al., 2005 (link)). The same study found that a maximum plasma concentration of 5-HMF at 30 min after infusion for doses of 100 mg/kg, therefore by initiating hypoxia at 20 min post infusion ensures that the maximum plasma concentration is achieved during hypoxia. Six animals were assigned to each group. Animals were subjected to 30 min challenge of 15% hypoxia, then to 30 min challenge of 10% hypoxia, and lastly to 30 min challenge of 5% O₂ hypoxia. Cardiac function measurements were taken 25 min into respective hypoxic stage. Representation of experimental timeline is presented in Figure 1B.

LPS (Escherichia coli O111:B4) and 5-HMF were purchased from Sigma-Aldrich (St. Louis, MO, USA). Dulbecco’s Modified Eagle Medium (DMEM) was purchased from Gibco (ThermoFisher, Gaisburg, MD, USA). Fetal bovine serum (FBS) was purchased from Lonsera (Lonsa Science SRL, Guichon, Uruguay). MTT cell proliferation and cytotoxicity assay kit was purchased from Phygene life sciences (Fuzhou, China). The primary rabbit monoclonal antibodies against p38 MAPK (D13E1) XP^®, SAPK/JNK Antibody, p44/42 MAPK (ERK1/2) (137F5), phospho-p38 MAPK (Thr180/Tyr182) (D3F9) XP^®, phospho-SAPK/JNK (Thr183/Tyr185) (81E11), phospho-p44/42 MAPK (ERK1/2) (Thr202/Tyr204) (D13.14.4E) XP^®, Akt (pan) (C67E7), mTOR (7C10), phospho-Akt(Ser473) (D9E) XP^®, phospho-mTOR (Ser2448) (D9C2) XP^®, IκBα (44D4), NF-κB p65 (D14E12) XP^®, phospho-IκBα (Ser32) (14D4) and phospho-NF-κB p65 (Ser536) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Standards for AA, 5-HMF, ¹³C₃-AA, ¹³C₆-HMF, 3-deoxyglucosone, quinoxaline, 2-methylquinoxaline, o-phenylenediamine (OPD),glucose and frucose were purchased from Sigma-Aldrich (St. Louis, MO, USA.). L-asparagine analytical standard was purchased from Beijing Solarbio Technology Co., Ltd (Beijing, China). HPLC-grade methanol and acetonitrile were obtained from Merck Company (Darmstadt, Germany). HPLC-grade formic acid was obtained from Shanghai Anpel Technology Co., Ltd (Shanghai, China). Distilled water was purchased from Watsons Water Co., Ltd. (Guangzhou, China). HPLC-grade n-heptane, D-glucose, asparagine and linoleic acid were obtained from Aladdin Biochemical Technology Co. Ltd. (Shanghai, China). O-phthalaldehyde (OPA) and 2-mercaptoethanol (2-ME) were purchased from Macklin Biochemical Technology Co., Ltd (Shanghai, China).

The sweet chestnut bark was obtained from a local company Tannin Sevica (Slovenia). Gallic, ellagic and levulinic acids, sugars, 5-HMF, furfural, Na₂CO₃ and phenol were obtained from Sigma-Aldrich (Steinheim, Germany). Folin-Dennis and Folin-Ciocalteu reagents and sulfuric acid (95–97%) were obtained from Merck (Darmstadt, Germany). All other chemicals used for HPLC were of analytical grade.