# Lichens as a Natural Source of Compounds Active on Microorganisms of Human Health Interest
## 3 Results
### Fractionation of lichen Extracts  
From the chromatographic separation of LC1 and LC2, 19 and 18 fractions were obtained, respectively. Among the 19 samples of LC1, TLC analysis showed organic compounds only in fractions 5-17 (Figure 2); on the contrary, LC2 fractions did not exhibit a clear composition. Nevertheless, both LC1 and LC2 fractions were analyzed for antimicrobial activity through the agar well diffusion assay as previously described. Results obtained for LC1 fractions showed a good antimicrobial activity in samples 8-16 toward all indicator strains, although with a reduced diameter of the inhibition zone, compared to the total LC1 extract, probably due to the fractionation of the active compound into the various samples. On the contrary, in all LC2 fractions, no antimicrobial activity was detected; therefore, the  
\begin{table}
\begin{tabular}{c c c c c c c c c} \hline  & \multicolumn{4}{c}{**LC1**} & \multicolumn{4}{c}{**LC2**} \\ \hline  & **1 d** & **7 d** & **15 d** & **30 d** & **1 d** & **7 d** & **15 d** & **30 d** \\ \hline _Staphylococcus aureus_ ATCC 6538 & 8 \(\pm\) 2 & 4 \(\pm\) 0.5 a  & 0 b  & 0 b  & 0 b  \\ _Enterococcus faecalis_ ATCC 29212 & 24 \(\pm\) 2 & 24 \(\pm\) 1 & 0 d  & 0 d  & 24 \(\pm\) 0.5 & 22 \(\pm\) 1 & 0 d  & 0 d  \\ _Bacillus subtilis_ ATCC 6633 & 16 \(\pm\) 2 & 15 \(\pm\) 1 & 0 c  & 0 c  & 16 \(\pm\) 0.5 & 14 \(\pm\) 1 & 0 c  & 0 c  \\ _Escherichia coli_ ATCC 8739 & 24 \(\pm\) 0.5 & 0 d  & 0 d  & 24 \(\pm\) 0.5 & 0 d  & 0 d  & 0 d  \\ _Pseudomonas aeruginosa_ ATCC 27853 & 22 \(\pm\) 2 & 0 d  & 0 d  & 22 \(\pm\) 1 & 0 d  & 0 d  & 0 d  \\ _Burkholderia cepacia_ MM 08 & 6 \(\pm\) 1 & 0 d  & 0 d  & 6 \(\pm\) 2 & 0 d  & 0 d  & 0 d  \\ _Klebsiella pneumoniae_ MM 01 & 12 \(\pm\) 0.5 & 0 d  & 0 d  & 12 \(\pm\) 2.5 & 0 d  & 0 d  & 0 d  \\ _Candidida albicans_ ATCC 10231 & 40 \(\pm\) 2 & 40 \(\pm\) 3 & 39 \(\pm\) 0.5 & 38 \(\pm\) 3 & 40 \(\pm\) 1 & 40 \(\pm\) 1 & 37 \(\pm\) 2 & 37 \(\pm\) 1 \\ \hline \end{tabular}
\end{table}
Table 1: Antimicrobial activity of cyclohexane (LC1) and acetone (LC2) lichen extracts against human pathogens using the agar well diffusion assay. Data are expressed in mm (diameter of inhibition zone) detected 1, 7, 15, and 30 days (d) after extraction procedure. Values are the arithmetic mean (n = 3) \(\pm\) SD (standard deviation). Values marked with letters were statistically different according to t-test and ANOVA; \(p\) < 0.05 (*), \(p\) < 0.01 (*), \(p\) < 0.001 (*), and \(p\) < 0.0001 (*).  
Figure 1: Antimicrobial activity of LC1 and LC2 lichen extracts against (**a**) _Enterococcus faecalis_ ATCC 29212 and (**b**) _Candidida albicans_ ATCC 10231 7 and 30 days, respectively, after the extraction procedure.  
Chromatograms of samples 8-16 obtained from LC1 fractionation revealed the presence of a significant peak at \(\rm t_{R}\) 10.8 min (Figure 6). Liquid chromatography/mass spectrometry (LC-MS) analysis, in accordance with the GNPS database [42], showed that this compound was traceable to a scabrosin (MW = 532.6) (Figure 7) derivative that was absent in the remaining fractions of LC1, as well as in all the LC2 samples. These results suggest that the antimicrobial activity can most be likely associated with the scabrosin derivative that generates the peak at 10.8 min.