Spinach ( Spinacia oleracea L . ) is a green herbaceous annual leafy vegetable widely cultivated. It is considered a functional food for its nutritional composition, phytochemicals and bioactive compounds that contribute to reduce oxidative stress, inducing the secretion of satiety hormones helping to promote protection mechanisms against hypoglycaemia, cancer and obesity. In this study spinach leaves were treated in order to extract the protein fraction ribulose‐1,5‐bisphosphate carboxylase/oxygenase (RuBisCO) by using calcium chloride (CaCl 2 ) to improve the removal of pigments in the extract. Functional properties, free amino acids content determined after derivatization with fluorenylmethyloxycarbonyl (Fmoc) group by the HPLC‐DAD technique, together with phenolic and flavonoid contents (TPC and TFC, respectively), antioxidant and enzyme inhibitory assays, were assessed in both samples treated with (e.g., S2) or without (e.g., S1) CaCl 2 . Two independent replicates of each sample were produced following the same procedure. Around 20 mg of each sample has been used for characterization. Data reveal that the sample S1 contains lysine (0.040 mg/mg DE) as principal amino acid while it is absent in S2. Both extracts present a comparable TPC content; conversely, the TFC is higher in the S1 sample (3.65 mg RE/g) than that in S2. Biological assays reveal higher antioxidant activity by ABTS (21.35 mg TE/g) and metal chelating (32.30 EDTAE/g) assays for S1 than sample S2. Both samples show similar inhibition of tyrosinase (47.45 mg KAE/g and 49.90 mg KAE/g for S1 and S2, respectively) and glucosidase enzyme (8.15 mmol ACAE/g and 8.06 mmol ACAE/g for S1 and S2, respectively), prompting further studies in developing more efficacious protein extraction methods for the design of enriched foods and beverages.
Marinaccio et al. (2026) studied this question.