Biological Samples

Pang et al. [116] fabricated a polymer monolith column with 1-vinyl-3-hexylimidazolium bromide ([ViC6MIM][Br]) IL which was used for the on-line SPE isolation of betamethasone, norgestrel, halcinonide, beclomethasone dipropionate and testosterone propionate from human plasma. The developed SPE-HPLC-UV method o ffered the e ffective extraction of the analytes (93–105%), which allowed the target compounds to be quantified with LODs of 1–2 ng/mL. In a study by Liu et al. [117] a poly(ionic liquid-glycidylmethacrylate-coethyleneglycol dimethacrylate) (IL-GMA-co-EDMA) monolithic column with 1-vinyl-3-butylimidazolium chloride ([ViC4MIM][Cl]) was synthesized and applied as an SPE sorbent in the on-line SPE-HPLC-UV method for the determination of nifedipine, nitrendipine and felodipine in human plasma samples. The best extraction of the analytes and purification of the matrix sample was obtained when a methanol/water mixture was used as the eluting agent. It allowed the three antihypertensive drugs in human plasma samples to be determined with LODs of 2–3 ng/mL. Ferreira et al. [118] used 1-vinyl imidazole and 1,4-butane-sultane to create a silica-anchored IL-based material which was applied as a sorbent in an SPE system coupled online with HPLC-MS/MS for the quantification of the antibiotic ceftiofur in bovine milk samples. The extraction e fficiency ranged from 70 to 130%, and the LOD was 0.1 μg/L. A sol–gel synthesis of three hybrid materials containing [C4MIM][PF6], [C6MIM][PF6] and [C8MIM][PF6], attached by covalent bonds, was published by da Silva and Mauro Lanças [119]. These IL-based hybrid materials were applied as the sorbents in o ff-line SPE for the isolation of five sulfonamides and trimethoprim from bovine milk samples. The results indicated that the extraction e fficiency of the analytes systematically decreased when the alkyl chain of the IL increased from C4 to C8. This was probably caused by the reduction of the electron density and the steric hindrance from the methyl group on the three-substituent site of imidazole rings, which weakened the π–π interaction between the electron-rich benzene ring of the target compounds and the imidazole rings of the used ILs. The best e fficiency was o ffered by an IL (C4)-based sorbent which was applied for the isolation and preconcentration of sulfonamide in bovine milk by the on-line SPE-HPLC-MS method. The LODs for the method developed were in the range of 1.5–2.25 μg/mL, with extraction recoveries from 74 to 93%. Yan et al. [120] developed modified dummy molecularly imprinted microspheres (DMIMs) based on [AC2MIM][Br] as the co-functional monomer and phenylephrine as the dummy template. These DMIMs were used as the SPE sorbent for the isolation of clenbuterol and clorprenaline from urine samples. The obtained results confirmed that they were able to more e ffectively extract the analytes and remove the matrix interferences than with other tested commercial sorbents such as HLB, PCX, C18 and SCX. For the DMIMs, the extraction e fficiency ranged from 93.3 to 106%. The developed DMIMs-SPE-HPLC method allowed the analytes to be quantified with LODs of 0.19 and 0.070 μg/<sup>L</sup> for clorprenaline and clenbuterol, respectively. Ma and Row [1] synthesized a molecularly imprinted monolithic column using levofloxacin and ciprofloxacin as templates, 1-vinyl-3-ethylimidazolium bromide ([ViC2MIM][Br] as the functional monomer, and graphene oxide (GO) as the core material. When the e fficiency of the IL-based imprinted monolithic column was tested as the SPE sorbent for the extraction of levofloxacin and ciprofloxacin from human urine, the best results were achieved using water as the washing agent, and a mixture of ethanol/acetic acid (7:3 *v*/*v*) for the elution of the analytes. The main advantages of the developed SPE protocol were the e ffective purification of the matrix sample, and the good extraction recovery of the analytes (89.5% and 92.5% for levofloxacin and ciprofloxacin, respectively). However, relatively low sensitivity of the developed SPE-HPLC-UV method was also observed (LODs from 0.06 to 0.27 μg/mL). Wu et al. [121] used an SPE procedure based on hemimicelles and admicelles (mixed hemimicelles) supported by an IL for the simultaneous extraction of five cephalosporins from biological samples. In this technique, the sorbent possesses adsorbed ionic surfactants on the surface of mineral oxides (e.g., SDS or IL) which enables two mechanisms to occur for the retention of the analytes—hydrophobic and electrostatic interactions. In e ffect, the extraction e fficiency can be improved. The authors tested seven di fferent surfactants, such as SDS, cetyltrimethylammonium bromide (CTAB), [ViC6MIM][Br], [C4MIM][Br], [C12MIM][PF6], [C16MIM][Br] and [C12MIM][Br]. The best recoveries were obtained for the long-chain IL [C16MIM][Br], which confirms data presented in (Figure 4).

The imidazolium-based IL with a longer alkyl side chain was probably able to strengthen the directionality of hydrogen bonds and van der Waals forces. In consequence, the interactions between the mixed hemimicelles and the hydrophobic regions of target compounds were more intensive and the efficiency increased. Taghvimi et al. [122] prepared mixed hemimicelle magnetic dispersive solid-phase extraction (MHMDSPE) based on carbon-coated magnetic nanoparticles and supported by the IL (IL-C/MNPs) for the extraction of tramadol from urine samples. In this study, MHMDSPE conditions were optimized, including both the selection of the adsorbent type and the solvent used as a desorbing agent. The results indicated that the IL-C/MNPs with [C6MIM][PF6] was more effective than that based on Fe3O4 NPs.

**Figure 4.** Comparison of the types of surfactants on the extraction e fficiency of cefoperazone and cefotaxime. Figure adopted from the reference [121] with copyright permission.

This was probably related to the presence of carboxyl and hydroxyl groups on the surface of IL-C/MNPs, which improved the dispersion of the magnetic nano-adsorbent in the urine medium. In e ffect, stronger interactions between the analyte and the magnetic nano-adsorbent occurred, which improved the extraction efficiency. The best desorbing solvent was acetone, which allowed a recovery of 94% to be obtained. Yan et al. [123] prepared IL-modified magnetic polymer microspheres (ILMPM) based on Fe3O4 NPs and [C4MIM][PF6] used as a magnetic adsorbent of MDSPE for the determination of sulfamonomethoxine sodium and sulfachloropyrazine sodium in urine samples. The developed ILMPM-SPE sorbent provided a higher purification ability and extraction recovery of the tested analytes compared with magnetic polymers based on using 4-vinyl pyridine, methacrylic acid and acrylamide as monomers. A report was also published describing matrix solid-phase dispersion coupled with homogeneous ionic liquid microextraction (MSPE-HILME) applied for the extraction of sulfamerazine, sulfathiazole, sulfamethazine, sulfadoxine, sulfachloropyridazine, sulfaphenazole and sulfisoxazole from animal tissues [124]. In the study, three kinds of hydrophilic ILs, including [C4MIM][BF4], [C6MIM][BF4], and [C8MIM][BF4] were tested in MSPD and HILME simultaneously. The results confirmed that higher extraction recoveries of the analytes were obtained with the C4 IL than those observed with C6 and C8 ILs. This was related to the significant loss of C6 and C8 ILs in MSPD, which resulted in a small volume of the IL phase and low extraction yields of the target analytes. Compared to C6 and C8 ILs, the C4 IL possesses higher water miscibility and lower viscosity, which facilitates the transfer of target analytes from the sample matrix to the extraction solvent. In this study, this effect was predominant in respect to the extraction capacity of the IL, which often increases with the increase in the alkyl chain length of the IL [125]. Finally, water was selected as the elution solvent in MSPD because of the more effective extraction of sulfonamides, which are water-soluble polar compounds. In this procedure, the C4 IL was mixed with the dispersant and the sample before introduction to the MSPD column, and the IL phase was collected after HILM. When the MSPD-HILME method was coupled to HPLC-UV, the recoveries of the sulfonamides ranged from 85.4 to 118.0%. The LODs for the analytes were 4.3–13.4 g/kg. The application of magnetic core-shell nanoparticles (mag-NPs) of SiO2@Fe3O4 type, covalently modified with the IL (dimethyl octadecyl [3-(trimethoxysilyl propyl)]ammonium chloride) as the MSPE material for the extraction of tolmetin, indometacin and naproxen from blood samples was also described in the literature [126]. The synthesized mag-NPs were applied as the adsorbent in MSPE according to the protocol presented in Figure 5.

**Figure 5.** Schematic illustration of extraction procedure for tolmetin (TOL), indomethacin (IND) and naproxen (NAP) from blood samples. Figure adopted from the reference [126] with copyright permission.

The results of the study showed that the IL addition provided a more effective extraction of the NSAIDs probably due to an increase in both hydrophobic and π-π dipole or electrostatic interactions between the adsorbent surface and the analytes. On the other hand, the adsorption of the cationic molecules onto the sorbent was limited because of the repulsion interaction with the adsorbent surface. In consequence, a better purification of the sample was also achieved. The optimized MSPE was coupled to HPLC-UV and used alone or after supercritical fluid extraction (SFE) before HPLC separation. These protocols resulted in LODs between 0.1 and 0.3 μg/<sup>L</sup> for MSPE-HPLC and 0.2 to 0.3 mg/kg for SFE-MSPE-HPLC, respectively.
