Hen egg white lysozyme (HEWL) was hydrolyzed with papain, trypsin and a combination of the two to isolate antioxidant peptides. The prepared hydrolysates were evaluated for antioxidant activity using DPPH and ABTS radical scavenging, metal ion chelation and lipid peroxidation inhibition. The obtained hydrolysate by a combination of the two enzymes exhibited the highest antioxidant activity compared to other hydrolysates and elected for isolation of antioxidant peptides by reverse-phase high-performance liquid chromatography (RP-HPLC). A most potent fraction namely F2 fraction, identified to be NTDGSTDYGILQ-INSR (MW: 1753.98 ± 0.5 Da) using tandem mass spectrometry. The antimicrobial activity of the F2 peptide was tested using radial diffusion assay (RDA). Our results showed that this peptide has inhibitory effects on both Gram-negative and Gram-positive bacteria. Minimum inhibition concentration (MIC) values of the F2 peptide against Escherichia coli and Leuconostoc mesenteroides bacteria were 355.64 (±2.2) and 442.25 (±2.8) lg/ml, respectively.

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The binding of ciprofloxacin to human serum albumin (HSA) in the presence and absence of silver nanoparticles of three sizes was investigated for the first time. For this purpose fluorescence spectroscopy, circular dichroism, UV-vis spectroscopy, and ζ potential techniques were employed under physiological conditions. The titration results indicated that ciprofloxacin quenched the fluorescence intensity of HSA through a static mechanism, but in the presence of Ag nanoparticles of sizes I and II there were two different kinds of interaction behavior. Results of circular dichroism indicated that the secondary structure of HSA was modified with increasing ciprofloxacin concentration. A comparison between resonance light scattering of binary and ternary systems allowed us to estimate the effect of silver nanoparticles on the initial formation and aggregation of ciprofloxacin with HSA. The ζ-potential results suggested induced conformational changes on HSA, thus confirming the experimental and theoretical results.

In this study, two novel antimicrobial peptides from the skin secretions of the marsh frog, Rana ridibunda, named temporin-Ra and temporin-Rb, were identified and purified using RP-HPLC. Temporin-Ra and temporin-Rb are composed of 14 and 12 amino acids, respectively. Our results show that these peptides have inhibitory effects on both gram-negative and gram-positive bacteria, especially antibiotic resistant strains prevalent in hospitals, such as Staphylococcus aureus and Streptococcus agalactiae. The sequences and molecular weights of these peptides were determined using tandem MS. The molecular masses were found to be 1242.5 Da for temporin-Rb and 1585.1 Da for temporin-Ra. Human red blood cells tolerated well exposure to temporin-Ra and temporin-Rb, which, at a concentration of 60 µg/ml, induced 1.3% and 1.1% hemolysis, respectively. MIC values of these peptides are suitable for potent antimicrobial peptides. The low hemolytic effect and wide-spectrum antimicrobial activity suggest a possible therapeutic application of these novel peptides.