## 4 Molecular Imprinted Polymer in Solid Phase Extraction (MISPE)
### Offline Modes  
In recent years, offline MISPE has seen a rise in popularity. Offline MISPE operations are identical to traditional SPE processes consisting of four important steps (conditioning, sample loading, washing, and elution). The MIPs are packaged into cartridges using a standard process that contains 15-500 mg of MIPs [64]. Firstly, the cartridge is conditioned with a conditioning solvent in order to optimize interactions between the analyte's target in the sample and the MIP [96]. Sample loading is  
Figure 14: MISPE procedure [95]. Figure 13: (a) FT-IR spectra collected for the MIP1 product (synthesized with MAA as functional monomer and EGDMA as crosslinker); (b) FT-IR spectra collected for the MIP7 product (synthesized with 4 VP as functional monomer and EGDMA as crosslinker) [61]. one of the most essential steps of MISPE. In this step, the sample medium must be carefully chosen because it has a significant effect on the MIP's recognition towards the analyte. The type of template/monomer interactions that occur during polymerization, as well as the porogen used, determines the loading solvent. To bind analytes at the imprinted locations, an electrostatic driving force, such as ionic interactions and hydrogen bonding, is typically used. Therefore, when loading samples, a low-polarity solvent should be used to establish the interaction towards analytes. Toluene, chloroform, acetonitrile, and dichloromethane have been reported as the most widely used solvents. However, it should be highlighted that when an aqueous solution containing the analyte of interest is saturated at the MIP surface, hydrophobic interactions might cause the analyte and other interfering chemicals to be retained non-selectively together with the analyte [55]. The washing step is utilized to maximize the particular interactions between the analytes and MIP while removing the interfering components that have been retained in the polymer matrix. The washing solvent should be selected based on the analyte, impurity, and the characteristics of the components. This step usually uses a low-polarity organic solvent such as dichloromethane, chloroform, toluene, or a mixture of these solvents. When electrostatic interactions drive recognition, the usage of porogens during polymerization, similar to the loading step, is a better option. However, using an aqueous solution on a MIP column is difficult since many porogens are incompatible with water. Thus, the MIP column should be nitrogen-dried prior to the washing stage. The polarity of the solvent, on the other hand, must be enhanced when non-specific interactions are needed. To improve washing selectivity, a small amount of polar components such as water, alcohol, acetonitrile, or base is frequently utilized. In this case, it is necessary to manage the washing solvent volume, additive, and pH [55]. The final step in MISPE is elution. To achieve significant enrichment factors, only a small amount of solvent is necessary. \(\text{Protic/polar solvents}\) such as acetonitrile or water, or a mixture of both are widely applied in the elution step. Weak acids (acetic acid or trifluoroacetic acid) or weak bases (triethylamine or pyridine) in small amounts may be used to achieve significant recoveries due to the strong interactions between the MIP and the analyte. Following elution, the eluate typically undergoes a re-constitution step (dried, followed by dissolving in a suitable solvent) to increase the extraction enrichment factor. For the quantification of the analyte, several detection methods such as chromatography, capillary electrophoresis (CE), electrochemical methods, and atomic absorption spectrometry (AAS) is used. However, the offline approach typically takes a lengthy amount of time, which potentially increases analytical mistakes [55]. [PERSON] et al. 2012 [97] analyzed the compound Kirenol using the offline mode. Kirenol is a terpenoid derived from Siegesbeckiae, a Chinese herbal medicine that has been used to treat inflammatory, allergic, and arthritic conditions.