*3.5. Superoxide Anion Generation and Elastase Release Assays by Human Neutrophils*

Human neutrophils were obtained from the venous blood of healthy adult volunteers (20−30 years old), following a reported procedure [37]. Superoxide anion generation by fMLF (0.1 μM)/CB (1μM)-activated neutrophils was evaluated based on the reduction of ferricytochrome c, as previously described [37,40]. Elastase release by the fMLF (0.1 μM)/CB (0.5 μM)-activated neutrophils was determined using N-methoxysuccinyl-Ala-Ala-Pro-Val-p-nitroanilide as the elastase substrate, according to a previous protocol [37,40]. The concentration was 10 μM for compounds **1**–**9**. Genistein was used as a positive control.

#### *3.6. Cytotoxicity Test*

A lactate dehydrogenase (LDH) assay kit (Promega, Madison, WI, USA) was utilized to evaluate the cytotoxicity of the samples in human neutrophils. Human neutrophils were treated with DMSO or compounds **1**–**9** for 15 min at 37 ◦C. Cell-free supernatants were collected, and the amount of LDH was evaluated [37].
