## 2 Results and Discussion
### Method Validation
#### 2.1.5 Robustness  
The robustness of an analytical method embodies its ability to remain unaffected by the minor, deliberate variations of one or more parameters, demonstrating its reliability during normal usage [43]. In this case, the robustness was evaluated in terms of small but thought changes in the chromatographic parameters, i.e., column temperature (\(\pm\)5 \({}^{\circ}\)C), flow rate (\(\pm\)0.2 mL/min), mobile phase ratio (\(\pm\)2%) and wavelength (\(\pm\)2 nm) (Table 5). The separation between the two peaks is of no interest since their time gap is very large (\(\geq\)3.6 min). Thus, the separation is no issue regardless of the changes made to the chromatographic parameters. The peak areas of both compounds remained constant. The most significant difference was noticed in the asymmetry factor, especially for Dox, when lowering the column oven temperature, the flow, and the percentage of acetonitrile (ACN) in the mobile phase. These factors directly influence the column and compound interaction. No significant changes were observed in the Amox and Dox retention time in different conditions tested (RSD \(<\) 2%). Therefore, the obtained results indicated that the effects of the considered, deliberate variations on the chromatographic parameters were insignificant, and the proposed method was robust for its envisioned applications. Although several analytical techniques are available for the individual quantification of these antibiotics, their analysis is challenging due to their low levels and complex sample matrices. Liquid chromatography is one of the most used analytical techniques used in pharmaceutical analysis and drug control [56]. The validated HPLC-DAD method for  
\begin{table}
\begin{tabular}{c c c c c c c c c} \hline \hline  & & & \multicolumn{3}{c}{**Intraday (n = 5)**} & **RSD (\%)** & \multicolumn{3}{c}{**Interplay (n = 6)**} & **RSD (\%)** \\ \hline
**Analyte** & **Matrix** & \begin{tabular}{c} **Fortified** \\ **(\#g/g)** \\ \end{tabular}  & \ \begin{tabular}{c} **Measured** \\ **(\#g/g)** \\ \end{tabular}  & \begin{tabular}{c} **RSD (\%)** \\ \end{tabular}  & \multicolumn{3}{c}{**RSD (\%)**} \\ \hline Amox & & 30.0 & 30.3 \(\pm\) 0.34 & 1.12 & 101 & - & - & - \\  & & 60.0 & 60.4 \(\pm\) 0.72 & 1.19 & 101 & - & - & - \\ Dox & & 30.0 & 29.9 \(\pm\) 0.55 & 1.12 & 99.8 & - & - & - \\  & & 60.0 & 60.4 \(\pm\) 0.80 & 1.32 & 101 & - & - & - \\ Amox & Tablet & 20.0 & - & - & - & 20.1 \(\pm\) 0.65 & 3.23 & 100 \\  & & 35.0 & - & - & - & 35.3 \(\pm\) 0.86 & 2.44 & 101 \\ Dox & & 20.0 & - & - & - & 20.1 \(\pm\) 0.87 & 4.34 & 101 \\  & & 35.0 & - & - & - & 34.8 \(\pm\) 0.84 & 2.40 & 99.3 \\ \hline
**Analyte** & **Matrix** & \begin{tabular}{c} **Fortified** \\ **(\#g/mL)** \\ \end{tabular}  & \begin{tabular}{c} **Measured** \\ **(\#g/mL)** \\ \end{tabular}  & \begin{tabular}{c} **RSD (\%)** \\ \end{tabular}  & \begin{tabular}{c} **Accuracy (\%)** \\ \end{tabular}  & \begin{tabular}{c} **Measured** \\ **(\#g/mL)** \\ \end{tabular}  & \begin{tabular}{c} **RSD (\%)** \\ \end{tabular}  & \ \begin{tabular}{c} **RSD (\%)** \\ \end{tabular}  & \begin{tabular}{c} **Recovery (\%)** \\ \end{tabular}  \\ \hline Amox & & 20.0 & 20.2 \(\pm\) 0.37 & 1.82 & 100 & - & - & - \\  & & 50.0 & 49.9 \(\pm\) 0.60 & 1.20 & 101 & - & - & - \\ Dox & & 20.0 & 20.1 \(\pm\) 0.29 & 1.73 & 101 & - & - & - \\ Amox & Wastewater & 50.0 & 50.1 \(\pm\) 0.87 & 1.46 the simultaneous determination of Amox and Dox is simple, rapid, reliable and low cost considering its main advantages; namely, it requires no sample preparation and, thus, no expensive equipment, no expensive solvents (especially, it uses only ultrapure water), time-consuming steps; requires no buffers in the mobile phase; the mobile phase is easy to prepare; it uses only a binary pump and a DAD detector, which are, besides the most common type of pump and detector, the ones with the lower prices.