## 4 Discussion  
Despite the large amount of literature in the field of the optimization of GDL biosynthesis, there are no data on the separation and purification of this compound from the post-reaction mixtures. The published results are based only on the chromatographic determinations of the samples taken directly from individual culture stages. Considering the fact that the efficiency of the extraction, purification, and isolation of this compound is a key stage which generates significant costs in relation to the entire biotechnological cycle and is decisive for the application of the methods of biotechnological synthesis of GDL in industry, research was undertaken on how to effectively isolate this compound from reaction mixtures. Our focus was on extraction, hydrodistillation, and lactone adsorption. Liquid-liquid extraction is the most mature and commonly used method of separating fragrances compounds. Solvents commonly used in the extraction are dichloromethane, chloroform, ethyl acetate, diethyl ether, and \(n\)-hexane [36; 37]. They were also used (except for ethyl acetate) in GDL extraction. The best efficiency obtained in this study with the use of diethyl ether (99.9%) justifies its widespread use in the extraction of lactones from microbiological media [19; 24; 25; 26]. The similar efficiency and reproducibility of GDL extraction for dichloromethane and chloroform allow the conclusion that due to difficulties in extraction on a larger scale with diethyl ether (due to its high volatility and tendency to form explosive peroxides), it is possible to replace it with these solvents. The effectiveness of dichloromethane and chloroform is also confirmed in the literature. In the research on quantitative determination of \(N\)-acylhomoserine lactones, conducted by the team of [PERSON] et al. [38], the best extraction results with almost the same yields were obtained for the above-mentioned solvents. Canbay [37], conducting liquid-liquid extraction of the volatile compounds from rose aromatic water, obtained complete extraction for chloroform, dichloromethane, and ethyl acetate. \(n\)-Hexane showed the worst extraction parameters, similar to our GDL extraction study. The effectiveness of dichloromethane in the extraction of terpenoids from aqueous solutions was also confirmed by the team of [PERSON] et al. [39]. The role of the volume of the solvent used in the extraction is highlighted by [PERSON] et al. [40], who conducted research on determination of lactones in milk fat. The authors conducted their study with the use of 0.5 to 1.5 mL of the solvent per 0.5 g of the extracted  
Figure 4: Desorption of the gamma-decalactone from adsorbents (zeolite, vermiculite, and Amberlite XAD-4) using five cycles of extraction with ethyl alcohol. sample. The results showed that the use of a large amount of an organic solvent for the extraction of lactone clearly improves the recovery rate. This conclusion is consistent with the data obtained in this study, where solvent-to-sample ratios of 1:2 or 1:1 (_v:v_) were used for GDL extraction. The difference in the extraction efficiency for the solvents used in these two volume variants was from 9 to 17%. [PERSON] et al. [40] showed that with a small amount of an organic solvent relative to the sample, long-chain lactones (above C\({}_{6}\)) are captured by triacylglycerlecrols that are present in the samples due to their hydrophobic nature. When a larger volume of solvent is used for extraction, the lactones are not affected by the triacylglycerglycerol matrix effect. Since the biotechnological synthesis of GDL uses an oil-in-water emulsion base (addition of 10 g/L castor oil as a precursor to GDL), it is highly probable that the lactone partially dissolves in the triglycerides of the unreacted oil. This may also explain the poorer reproducibility of the extraction results for the samples extracted with a 1:2 solvent-to-sample ratio (higher standard deviations).