Alcl3

For this experimental study, AlCl₃ , CdCl₂ , PbCl₄ , naringin, kaempferol, and quercetin were obtained from Merck (Darmstadt, Germany). Rutin, catechin and the remainder of chemicals and reagents used in this research were purchased from SigmaAldrich (St. Louis, MO, USA).

TFC was determined using a method being based on the interaction of flavonoids with AlCl₃ (Merck Chemicals, Darmstadt, Germany) that leads to the formation of a complex that can be determined via a UV/Vis spectrophotometer (BioTek Instruments, Winooski, VT, USA) at 420 nm [54 (link)]. The TFC values are expressed as mg rutin (Cayman Chemical Co., MI, USA) equivalents (RE) per gram of suspension based on a calibration curve with rutin (10–100 µg/mL). In addition, results were expressed as relative extraction efficacy (%) and compared to the nonprocessed bulk material.

Assay for the content of flavonoids in the fungal extracts was performed by spectrophotometric method with aluminum chloride according to Ghasemzadeh et al. [12 (link)]. In brief, fungal homogenate (1 mL), 5% sodium nitrite (NaNO₂, 0.7 mL, Merck KGaA) and 30% ethanol (10 mL) were mixed for 5 min, and then 10% aluminum chloride (AlCl₃, 0.7 mL, Merck KGaA) was added and mixed altogether. After 6 min, 1 mol/L sodium hydroxide (NaOH, 5 mL, Merck KGaA) was added, the mixture was diluted to 25 mL with 30% ethanol and let to stand for 10 min. The absorbance of the solution was then measured at 430 nm with a spectrophotometer (Shimadzu, Kyoto, Japan). A standard curve was plotted using quercetin. The assay was performed in triplicate.

The method previously described was used for determination of total phenolic content [15 (link)]. Accordingly, 20 μL of properly diluted BCFE sample were mixed with Folin–Ciocalteu reagent (Sigma) and Na₂CO₃ (20%) (Riedel de Hean). Then, samples were incubated at 45°C for 30 minutes. At the end of this period, the absorbance was measured at 765 nm by UV-Vis spectrophotometer (Thermo, Evolution 300). Results were expressed as mg gallic acid equivalents (GAE) per gram of dried extract.
For the determination of total flavonoid content, 500 μL of properly diluted BCFE sample were mixed with 10% AlCl₃ (Merck) and 1 M sodium acetate (Riedel de Hean). Following the incubation period (30 minutes at room temperature), the absorbance was recorded at 415 nm. Results were expressed as mg quercetin equivalents (QE) per g of dried extract [13 (link)].
The total proanthocyanidin content of the BCFE sample was measured by adding of 2.5 mL of vanillin (1%) (Fluka) and 2.5 mL of 9 M HCl (Sigma) in methanol to properly diluted extracts. After incubating at 30°C for 20 min, the absorbance was measured at 500 nm. Total proanthocyanidin content of samples was expressed as mg epigallocatechingallate equivalents (EGCG-E) per g of dry extract [13 (link)].

¹H and ¹³C NMR spectra were acquired using an Avance Bruker 300 spectrophotometer (Bruker Biospin, Mannheim, Germany), operating at the frequencies of 300 MHz for hydrogen and 75 MHz for carbon. The spectra were recorded in acetone-d6 solvent (Merck Group, Darmstadt, Germany). Mass spectral data were obtained using ultra-high-pressure liquid chromatography (UPLC) coupled to a quadrupole time-of-flight (QTOF) mass spectrometer detector, specifically a Shimadzu LCMS-9030 (Kyoto, Japan), via direct injection. The positions of O-CH₃ and OH groups were determined using UV-VIS spectroscopy within the range from 230 to 500 nm, conducted on a Jenway 6405 UV-VIS spectrophotometer (Staffordshire, UK). This analysis utilized displacement reagents, including AlCl₃, HCl, AcONa, MeONa, and H₃BO₃ (Merck Group, Darmstadt, Germany).

Nano Novin Polymer Co. (Sari, Iran) provided wet BCNF sheets for the study. From the local market, purple onion was purchased (Bandar Abbas, Iran).
Salts of metal, including Pb(NO₃)₂, KNO₃, Cu(NO₃)₂, Hg(NO₃)₂, CrCl₃, AlCl₃, NiCl₂ and MnSO₄, were supplied from Merck. Suitable amounts of these salts were dissolved in distilled water to prepare stock aqueous solutions. At first, the aluminium (III) chloride stock solution was prepared at 1000 ppm, followed by serial dilutions (100–1000 ppm). Acetate buffer solutions were prepared in the laboratory using sodium acetate and glacial acetic acid. The rest of the reagents (analytical grade) were provided from Sigma‐Aldrich.