## 2 Materials and Methods
### Antimicrobial Activity  
The antimicrobial potential of the phenolic extracts, derived from the pruning firewood and stem samples of the three chosen varieties, was assessed using the disk diffusion  
\begin{table}
\begin{tabular}{c c c} \hline
**Bacterial Species** & **Strains Designation** & **Source** \\ \hline _Escherichia coli_ & ATCC 25922 & American Type Culture Collection (control) \\ \hline _Acinetobacter baumannii_ & MJMC 525 & Trochenteric ulcer \\ \hline _Acinetobacter baumannii_ & MJMC 561 & Abscess \\ \hline _Pseudomonas aeruginosa_ & MJMC 526 & Sacred ulcer \\ \hline _Pseudomonas aeruginosa_ & MJMC 553 & Calcaneal ulcer \\ \hline _Pseudomonas aeruginosa_ & MJMC 559 & Chronic left leg ulcer \\ \hline _Staphylococcus aureus_ & ATCC 25923 & American Type Culture Collection (control) \\ \hline MRSA & MJMC 534-B & Sacred ulcer \\ \hline MRSA & MJMC 583 & Ulcer \\ \hline MRSA, Methicillin-resistant _Staphylococcus aureus_; ATCC, American Type Culture Collection. \\ \hline \end{tabular}
\end{table}
Table 1: Bacterial isolates tested.  
method originally outlined by [PERSON] et al. in 1966, with certain adaptations incorporated [20].  
In summary, the process began with the preparation of a bacterial inoculum. This was achieved by introducing a colony isolated from pure strains into a 0.9% NaCl solution, with the turbidity meticulously adjusted to match the 0.5 McFarland standard units. This prepared inoculum was then carefully spread across Petri dishes with a diameter of 90 mm, each containing 20 mL of Mueller Hinton Agar.  
Following this, sterile paper discs measuring six mm in diameter were uniformly placed on the surface of the agar plate that had been previously seeded with the bacterial inoculum. These paper discs served as carriers to absorb 15 \(\upmu\)L of the polyphenolic extract, which was prepared in a 10% Dimethylsiloxane(DMSO) solution. This methodology allowed us to effectively assess the impact of the polyphenolic extracts on the growth of the bacterial strains under investigation, providing valuable insights into the potential antimicrobial properties of the extracts derived from the pruning firewood and stem samples.  
After the overnight incubation at 37 \({}^{\circ}\)C, the next step involved measuring the diameter of the clear inhibitory zones that had formed around the discs impregnated with the polyphenolic extracts. These measurements were conducted in mm. Each experiment included essential controls for comparison: a negative control consisting of 10 \(\upmu\)L of DMSO, and a positive control employing a standard commercial antibiotic, specifically gentamicin (10 \(\upmu\)g).  
The evaluation of antibacterial activity was carried out using the following equation:  
% RIZD = ((IZD sample \(-\) IZD negative control)/IZD antibiotic standard) \(\times\) 100%.  
In this equation:  
\"% RIZD\" represents the percentage of the relative diameter of the inhibition zone, measured in mm.  
'IZD sample' corresponds to the inhibition zone diameter for the tested sample.  
'IZD negative control' signifies the inhibition zone diameter for the negative control (10 \(\upmu\)L DMSO).  
'IZD antibiotic standard' represents the inhibition zone diameter for the positive control (standard commercial antibiotic gentamicin).  
This analytical approach, based on the equation, has been previously established in studies by [PERSON] et al. in 2020 [9]. Importantly, the equation considers and compensates for potential effects of the solvent (DMSO) other than water, ensuring that the assessment of antibacterial activity is accurate and unbiased.