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Additional techniques could be used to expand or complement the characterization of THEDES, like polarized optical microscopy (POM) that shows information
about the morphology of the eutectic mixture. The mixture is observed under polarized light, and the presence or absence of crystals is evaluated as an evidence of the
formation of a homogeneous and amorphous mixture (Silva et al. 2018; Aroso
et al. 2016).
Biological Characterization
• Biological characterization – for pharmaceutical applications, specific characterization methodologies should be performed, in particular the evaluation of the
stability of the THEDES for long periods of time, its response to adverse environments, the evaluation of dissolution rates (Aroso et al. 2015), their bioavailability through biopharmaceutics classification system (BCS) and
pharmacokinetic studies, and their toxicity.
For pharmaceutical substances, the Food and Drug Administration (FDA) provides a classification system named biopharmaceutics classification system (BCS)
that serves as a guideline to predict intestinal drug absorption; however, this system
restricts the simulations to the solubility and intestinal permeability (Duarte et al.
2017; Savjani et al. 2012). The parameters established by BCS described in the literature classifies an API in four different classes (Table 3.1), in which class I is high
soluble and high permeable, class II is low soluble and high permeable, class III is
high soluble and low permeable, and class IV is low soluble and low permeable
(Chavda et al. 2010; Duarte et al. 2017; Silva et al. 2018; Varma et al. 2004).
Solubility measurements can be performed in a buffer solution which is most adequate to the administration route of the THEDES. The maximum solubility is determined by saturating solutions under the same conditions. In the case of the
permeability, in studies performed with therapeutic deep eutectic systems, it is possible to use synthetic membranes to mimic the effect on tissue, for example, membranes of polyethersulfone (Duarte et al. 2017). These studies allow the relative
comparison of the permeability between the API and the THEDES. Also, permeability studies could be made by transwell assays that allow to generate cell layers
in vitro and reconstitute their microenvironment and measure the permeability of a
compound, by measuring the barrier integrity of the cell layer through transepithelial/transendothelial electrical resistance (TEER).
In vitro cytotoxicity tests are necessary mainly for applications for human consumption. This screening is based on assays that test cell viability (MTS, MTT,
Alamar Blue) and measure the IC 50 for THEDES and APIs, in different cell lines
depending on the application (Faggian et al. 2016; Hayyan et al. 2016; Mano et al.
2016). Other in vitro assays could be performed regarding the future application of
THEDES, such as antioxidant activity, apoptosis, and reactive oxygen species,
among others (Hayyan et al. 2015; Mbous et al. 2017a, b).
F. Santos and A. R. C. Duarte
Additional techniques could be used to expand or complement the characterization of THEDES, like polarized optical microscopy (POM) that shows information
about the morphology of the eutectic mixture. The mixture is observed under polarized light, and the presence or absence of crystals is evaluated as an evidence of the
formation of a homogeneous and amorphous mixture (Silva et al. 2018; Aroso
et al. 2016).
Biological Characterization
• Biological characterization – for pharmaceutical applications, specific characterization methodologies should be performed, in particular the evaluation of the
stability of the THEDES for long periods of time, its response to adverse environments, the evaluation of dissolution rates (Aroso et al. 2015), their bioavailability through biopharmaceutics classification system (BCS) and
pharmacokinetic studies, and their toxicity.
For pharmaceutical substances, the Food and Drug Administration (FDA) provides a classification system named biopharmaceutics classification system (BCS)
that serves as a guideline to predict intestinal drug absorption; however, this system
restricts the simulations to the solubility and intestinal permeability (Duarte et al.
2017; Savjani et al. 2012). The parameters established by BCS described in the literature classifies an API in four different classes (Table 3.1), in which class I is high
soluble and high permeable, class II is low soluble and high permeable, class III is
high soluble and low permeable, and class IV is low soluble and low permeable
(Chavda et al. 2010; Duarte et al. 2017; Silva et al. 2018; Varma et al. 2004).
Solubility measurements can be performed in a buffer solution which is most adequate to the administration route of the THEDES. The maximum solubility is determined by saturating solutions under the same conditions. In the case of the
permeability, in studies performed with therapeutic deep eutectic systems, it is possible to use synthetic membranes to mimic the effect on tissue, for example, membranes of polyethersulfone (Duarte et al. 2017). These studies allow the relative
comparison of the permeability between the API and the THEDES. Also, permeability studies could be made by transwell assays that allow to generate cell layers
in vitro and reconstitute their microenvironment and measure the permeability of a
compound, by measuring the barrier integrity of the cell layer through transepithelial/transendothelial electrical resistance (TEER).
In vitro cytotoxicity tests are necessary mainly for applications for human consumption. This screening is based on assays that test cell viability (MTS, MTT,
Alamar Blue) and measure the IC 50 for THEDES and APIs, in different cell lines
depending on the application (Faggian et al. 2016; Hayyan et al. 2016; Mano et al.
2016). Other in vitro assays could be performed regarding the future application of
THEDES, such as antioxidant activity, apoptosis, and reactive oxygen species,
among others (Hayyan et al. 2015; Mbous et al. 2017a, b).
F. Santos and A. R. C. Duarte