Ferric 3 chloride

The ferric reducing antioxidant potential (FRAP) reagent was prepared by mixing 0.3 M of acetate buffer (Sigma-Aldrich, USA), 10 mM of 2,4,6-tripyridyl-s-triazine (TPTZ) (Sigma-Aldrich, USA) and 10 mM of ferric (III) chloride (Sigma-Aldrich, USA) in a 10:1:1 ratio, respectively (19 ). The solution was kept warm at 37 °C in a water bath. The absorbance of the FRAP solution as the blank was measured at 593 nm using a UV/VIS-spectrophotometer (Schott UVLine 9400, USA). Then, the reading of MTJ extract was obtained at the same wavelength. 8-tetramethyl-chroman-2-carboxylic acid (Trolox) (Sigma-Aldrich, USA) was used as the standard for the construction of the calibration curve. The result was expressed in millimolar Trolox equivalence per 100 g of MTJ (mM TEQ/100g). This assay was done in triplicates.

The edible insects were sourced from different provinces of South Africa: Mashonzha (G. belina) and Madzhulu (M. subhylanus) were sourced in the Vhembe district, Limpopo, and the black soldier fly larvae (H. illucens) was sourced from Cape Town, Western Cape, South Africa. The chemical reagents, 2,2 diphenyl-1-picrylhydrazyl (DPPH), 2,2′ azobis (2-methyl, 2,2-Azinobis (3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt (ABTS), Ferric (III) chloride, ethylenediaminetetraacetic acid (EDTA), tertiary butyl hydroquinone (TBHQ), ferrous (II) chloride and thiobarbituric acid (TBA) were obtained from Merk (Sigma-Aldrich, Kempton Park, South Africa). All the chemicals used in this study were of analytical grade, and chemical reagents were prepared according to standard analytical procedures. Prepared reagents were stored under conditions that prevented deterioration or contamination. The water used in the study was ultrapure water purified with a Milli-Q water purification system (Millipore, Microsep, Bellville, South Africa). The ethics committee of the faculty of applied sciences gave its approval to the study (215062965/05/2021).

Hydrocortisone (HYD), β-estradiol (E2), progesterone (PRO) and estrone (E1) were obtained from Sigma-Aldrich (St. Louis, MO, USA). The physical properties and chemical structure of the analytes are presented in Table S1. Sodium hydroxide, acetic acid (99.9%), methanol (HPLC grade), ferrous sulphate, ferric (III) chloride and β-cyclodextrin were all purchased from Sigma-Aldrich. A 20 mg L⁻¹ mixed hormone stock solution was prepared by dissolving an appropriate amount of the analyte in HPLC-grade methanol and was kept chilled at 2 °C. Standard solutions were prepared daily by diluting the stock solution with ultra-pure water (Direct-Q^® 3UV-R purifier system, Millipore, Merck).