## 2 Results and Discussion
### Optimization of Extraction Parameters
#### 2.1.2 Solid-Phase Extraction  
In the next stage of the experiments, the SPE efficiency of analytes for seven types of sorbents was estimated (Table 3). The results, presented in Table 4, indicate that the recovery (R) and potential for signal enhancement for DHPG, MHPG, and VMA after isolation from urine samples using the SPE technique depend on the physicochemical properties of the analytes, the type of sorbents, and the eluting/desorbing agents. For hydrophilic-lipophilic balanced (HLB) columns, where the sorbent is a hydrophillically modified styrene polymer, retention is primarily based on a reversed-phase mechanism. However, due to the hydrophilic modification, the sorbent is also prone to interactions with polar compounds. HLB sorbents offer good extraction recovery for non-polar to moderately polar acidic, neutral, and alkaline compounds, making them particularly suitable for the pretreatment of complex matrices such as blood, urine, and food. In our experiment with HLB columns, the highest recovery (R) was achieved using acetone as the eluent for DHPG (86.8 \(\pm\) 2.8%) and VMA (83.4 \(\pm\) 2.8%). For MHPG, the recovery was very low. Methanol (MeOH) proved to be a less effective eluent, with the best process efficiency obtained for the most hydrophobic compound, VMA, at 69.8 \(\pm\) 6.3%. The ACN:MeOH mixture was ineffective as an eluent for HLB cartridges, providing an extraction efficiency of less than 37.6 \(\pm\) 2.5%. \begin{table}
\begin{tabular}{c c c c c c c c} \hline \hline \multirow{2}{*}{**LLE Based Extraction**} & \multicolumn{2}{c}{**DHPG**} & \multicolumn{2}{c}{**MHPG**} & \multicolumn{2}{c}{**VMA**} \\ \cline{2-7}  & **R \(\pm\) SD (\%)** & **EF** & **R \(\pm\) SD (\%)** & **EF** & **R \(\pm\) SD (\%)** & **EF** \\ \hline \multirow{4}{*}{single process} & pH 4.6 & 159.4 \(\pm\) 10.5 & 1.6 & 11.6 \(\pm\) 1.3 & - & 103.8 \(\pm\) 5.5 & 1.0 \\ \cline{2-7}  & pH 5.5 & _no_ & - & 125.2 \(\pm\) 5.9 & 1.2 & 103.5 \(\pm\) 6.3 & 1.0 \\ \cline{2-7}  & pH 6.0 & 140.7 \(\pm\) 4.9 & 1.4 & _no_ & - & 197.8 \(\pm\) 5.7 & 1.9 \\ \cline{2-7}  & pH 9.2 & 151.4 \(\pm\) 6.0 & 1.5 & _no_ & - & 18.6 \(\pm\) 3.3 & - \\ \hline double process & pH 5.8 & 218.2 \(\pm\) 4.2 & 2.2 & 192.2 \(\pm\) 8.0 & 1.9 & 157.7 \(\pm\) 8.8 & 1.6 \\ \cline{2-7}  & pH 7.4 & 209.9 \(\pm\) 5.5 & 2.1 & _no_ & - & 166.7 \(\pm\) 4.2 & 1.6 \\ \hline \hline \end{tabular}
\end{table} 
Table 2: The results of average extraction efficiency (R) with SD value and enhancement factor (EF) for LLE based extraction procedures depending on pH of the sample matrix (n = 3). The octadecyl (C18) cartridges contain silica gel modified with octadecyl groups, enabling the extraction of non-polar compounds. In this case, the best extraction efficiency (R) values were achieved using methanol, which allowed enhancement factors (EF) of 3.0, 2.4, and 1.5 times for VMA, DHPG, and MHPG, respectively. The extraction results obtained by [PERSON] et al. also confirm high recovery for MHPG (81%) when C18 sorbent and a methanol-phosphate buffer mixture were used as the eluent [27]. Satisfactory results were also obtained with an eluting mixture of ACN:MeOH (50:50, \(v/v\)) for VMA and DHPG, where EF values were 1.9 times for VMA and 1.1 times for DHPG. Using acetone as the eluent, the best EF was achieved for VMA (1.5 times) and DHPG (1.2 times), while for MHPG, acetone was inefficient, yielding an extraction efficiency of 41.8 \(\pm\) 2.9% without