Antimicrobial and antioxidant activity of proteins from Feijoa sellowiana Berg. fruit before and after in vitro gastrointestinal digestion

Peptides and proteins are promising sources of antimicrobial and antioxidant compounds. Fruit are generally considered recalcitrant plant tissues for the difficulty to obtain high quality protein due to a low protein content and the presence of interfering substances such as polyphenols, carbohydrates, polysaccharides, pigments and starch. Pharmacological activity of Feijoa sellowiana is studied by our group from 90s demonstrating antibacterial, antifungal, antioxidant and anti-cancer activities of F. sellowiana acetonic extract, but so far no study on the protein component has been carried out. The aim of the present work has been the extraction of the proteins from F. sellowiana fruit and the study of their antimicrobial and antioxidant activity both in their native state and after in vitro gastrointestinal digestion, in order to obtain the formulation of nutraceutical or drug with antimicrobial and antioxidant activities to be administered per os. We developed for the first time a method for the extraction of the protein fraction from F. sellowiana fruit, which consisted of a mixture of proteins with molecular weight from 15 to over 66 kDa. The product of in vitro gastrointestinal digestion of F. sellowiana fruit proteins (digest) consisted instead of two peptides of apparent molecular weight of 5.61 and 2.04 kDa. We tested the bactericidal activity of these proteins against ATCC Gram-positive and Gram-negative bacteria. Clinical isolates (CI) of the same bacterial species were also used to compare their sensitivity to these proteins. Antimicrobial activity of protein fraction against the tested bacteria resulted surprising high, with MICs between 3.9 and 31.3 mg/ml for ATCC and 7.8 and 62.5 mg/ml for CI and with Minimum bactericidal concentrations (MBCs) between 15.7 and 62.6 mg/ml for ATCC and 31.3 and 125 mg/ml for CI. The Gram-negative bacteria were more sensitive, in particular Pseudomonas aeruginosa. Digest showed from 2 to 4 fold increased antimicrobial activity. MTT assays showed the nontoxicity of these proteins, both before and after digestion. These proteins also showed antioxidant activity which was increased 10 fold following the in vitro gastrointestinal digestion. Our results show that both antimicrobial and antioxidant activity resulted stronger respect to that previously obtained for the acetonic extract from the same fruit. The most significant result is the effectiveness of F. sellowiana protein fraction against P. aeruginosa, since infections due to this bacterium are particularly problematic because of their intrinsic resistance to multiple classes of antibiotics and their ability to acquire adaptive resistance during a therapeutic course. Moreover, the observation that the digest exhibits a higher antimicrobial activity respect to the undigested proteins is very important because AMPs often combine broad-spectrum antimicrobial activity with other interesting biological activities such as antitumor, immunostimulant, antiinflammatory, antiviral, etc. Moreover, in contrast to conventional antibiotics, AMPs are active on both dividing and quiescent cells including cells in biofilms. Finally, the absence of toxicity of F. sellowiana fruit extract and above all of the peptides obtained from the in vitro gastrointestinal digestion are important characteristics, which can suggest their potential use as a new class of antimicrobial drugs.