## 3 Results
### Optimization of the SB-MSPE Procedure  
A set of parameters were studied for the optimization of the SB-MSPE protocol in the order that is described in Figure 4. For example, when the mass of the adsorbent was examined, the type of adsorbent was powdered GO-Chm, with ACN as an elution solvent and 0.5 mL as the eluent volume, with the duration of the extraction and elution being 30 min and 15 min, respectively, while the stirring speed was kept at 500 rpm and the sample volume being 10 mL, with a pH value of 7 and 0% \(w/v\) addition of NaCl. Figure 3: UV spectra of (a) Alprazolam, ALP and (b) Flunitrazepam, FLT. Figure 2: Typical chromatograph of the two compounds in an aquatic stock solution after its sample pretreatment with the prepared nanocomposite GO-Chm. The first parameter that was examined was the type of adsorbent that would be used. Preliminary experiments were conducted on magnetic graphene oxide (mGO), magnetic graphene oxide with polystyrene (mGO-PS), magnetic activated carbon (BAX), magnetic graphene oxide with polyaniline (mGO-PANI) and on the nanocomposite, both in powder (GO-Chm) and bead (GO-Chm@SA) form. In Figure S1, there are the five chromatographs obtained in superposition, and in Figure 5 a bar diagram of normalized absolute recoveries is given. In mGO-PANI, ALP could not be determined, while the GO-Chm@SA beads showed a decline in their extraction efficiency when the percentage of the GO-Chm content was increasing. Thus, GO-Chm in the form of powder was used as the adsorbent for the extraction of the two BZs. The powdered GO-Chm nanocomposite was characterized by [PERSON] et al. (2016), with the XRD, SEM, VSM, DTG, DTA, FT-IR and XPS techniques. Through the XRD and VSM techniques, the mean size of the magnetic nanoparticles Fe\({}_{3}\)O\({}_{4}\) was calculated with the equation of Debye-Scherrer and found equal to 10.5 nm both in Chm and in GO-Chm, with a value of 9 emu/g for its saturated magnetization. The concentration of iron in the GO-Chm nanocomposite was approximately 12%, also having a rough surface, as indicated by SEM. Furthermore, with the DTG, DTA and FT-IR techniques, the identification of hydroxyl, carboxyl and amino groups was carried out, while the binding of chitosan onto the GO layer was confirmed. Figure 4: Methodological path of the optimization of the SB-MSPE procedure. The second parameter was the type and the volume of the elution solvent that would be used. ACN and MeOH were used in different volume ratios (100:0, 0:100, 70:30, 50:50, 30:70% \(v/v\)), while the possibility of adding a step of washing after the extraction and before the elution was studied with the addition of 1 mL of deionized water. The best results were shown with the pure ACN as a solvent, as it is a stronger eluent than MeOH, while the usage of deionized water showed a drop in the normalized absolute recoveries by 9-9.5% for the two analytes. Additionally, the elution volume that was selected was 0.5 mL, due to the increase in the preconcentration factor (or enrichment factor, EF) it provided. The prospect of using a nitrogen stream to evaporate the eluent and reconstitute it in a lower volume was also investigated, but the matrix effect would not allow their detection. Lastly, the EFs of the two compounds were calculated using the equation: \(\text{EF}=(\text{V}_{\text{sample}}/\text{V}_{\text{clement}})\times(\text{R}_ {\text{abs}}\%/100)\), where \(\text{R}_{\text{abs}}\) is the absolute recovery of each compound. In Figures 6 and 7, the bar diagrams of normalized absolute recoveries are demonstrated for the type and the volume of eluent, respectively. Figure 5: Bar diagram of the normalized absolute recoveries of the different adsorbents according to GO-Chm. Figure 6: Bar diagram of the normalized absolute recoveries of the different types of elution solvent according to acetonitrile (ACN). Another major parameter that affects the extraction efficiency is the mass of the adsorbent to be used.