## 3 Results  
### Phenolic Content of G. hederacea Extracts Obtained at Different Extraction Variants  
_G. hederacea_ CO\({}_{2}\) extracts obtained with different extraction conditions were tested for their biological activity. The yield of SC-CO\({}_{2}\) extraction with methanol as a co-solvent was established in the presented work at 9.58%. In SC-CO\({}_{2}\) extraction, density is extremely important and highly dependent on temperature and pressure variation [51; 52]. An increase in pressure favors an increase in density at a constant temperature, whereas an increase in temperature decreases the density at a constant pressure. In our study, we tested three temperatures (40, 50 and 60 \({}^{\circ}\)C) during the extraction process at constant pressure. At the lowest temperature we obtained the highest level of total phenolic content. As reported [PERSON] et al. (2020) [52] the increase in the temperature causes a greater intermolecular distance, and consequently the reduction of the solubility power of CO\({}_{2}\), by decreasing the density. In this study, the TPC in the obtained _G. hederacea_ extracts was analyzed spectrophotometrically using the Folin-Ciocalteu method. The results showed that the extract obtained in conditions of 40 \({}^{\circ}\)C/250 bar had the highest TPC value (138.33 \(\pm\) 5.00 mg GAE/g) compared with the samples obtained at temperatures of 50 \({}^{\circ}\)C and 60 \({}^{\circ}\)C (Table 1). The TPC values of the other two tested _G. hederacea_ extracts were lower than \({}^{1}/_{3}\) of the value obtained at 40 \({}^{\circ}\)C (43.00 \(\pm\) 3.04 mg GAE/g for 50 \({}^{\circ}\)C and 46.00 \(\pm\) 9.26 mg GAE/g for 60 \({}^{\circ}\)C). The most reported method for obtaining _G. hederaca_ extracts is aqueous extraction and the total phenolic content obtained in this study cannot be directly compared to those extracts, as different parameters (extraction method, time, pressure, solvent) had an influence on the final results. However, it is worth mentioning that [PERSON] et al. (2016) [32] reported total phenols from the aqueous _Gleduma_ extract in the range of 43.9 \(\pm\) 3.2-109.8 \(\pm\) 5.8 mg GAE/g, whereas [PERSON] et al. (2021) [53] reported an average phenolic content of 14.81 \(\pm\) 4.53 mg/g, and [PERSON] et al. (2012) [18] found a total phenol content of 79.70 \(\pm\) 0.193 mg GAE/g. The literature data indicate that SC-CO\({}_{2}\) can be used to obtain extracts rich in phenolic compounds [54]. However, due to the nonpolar nature of CO\({}_{2}\), some polar cosolvents such as ethanol, methanol or ethyl acetate need to be added to increase the extraction yield of phenolic compounds via the increase in solvation power [52; 55]. In the presented work, methanol was used as an example of a polar solvent due to its properties and costs. Moreover, the effect of temperature, pressure, flow rate and density of the supercritical CO\({}_{2}\) on the bioactive compound extraction process using different plant materials was analyzed previously [52]. According to some authors [51; 56], solvent density or pressure may influence the mass yield of the extract. According to a study by [PERSON] (2014), overall SFE yield increases with CO\({}_{2}\) density and co-solvent ratio [56]. [PERSON] et al. (2021) [51] reported that at 40 \({}^{\circ}\)C the solvent density affected the mass yields, whereas after increasing temperature to 50 \({}^{\circ}\)C, the solvent density did not influence yields, but rather affected the vapor pressure of the solute. Based on the obtained results, we can conclude that a temperature increase has a negative effect on phenolic solubility during extraction due to the decrease in the solvent's density. [PERSON] et al. (2009) [57] observed a decrease in the total phenolic compounds determined in an extract from _Gynura procumbens_ leaves when the temperature was increased in a conventional solvent extraction system.