# Microbial Biosynthesis of Lactones: Gaps and Opportunities towards Sustainable Production
## 4 Microbial Production
### Biotransformation
#### 4.1.2 Other Fungi
However, this attempt was unsuccessful due to growth defects of the engineered strain, probably due to the toxicity of ricinoleic acid. In the opposite way, the heterologous expression of oleate hydratase from _Stenetrophomonas maltolilia_ in _Escherichia coli_ led to a high-yield strain able to convert oleic acid into 10-hydroxysteric acid at 98% (50 g/L oleic acid resulted in 49 g/L of 10-hydroxysteric acid) [77]. This work was the basis for the development of one of the most successful biotransformation processes for the production of lactones reported so far [78]. The 10-hydroxysteric acid produced by the engineered _E. coli_ strain was used for the production of \(\gamma\)-dodecalactone by permeabilized whole cells of the oleaginous yeast _Waltomyces lipofer_[78; 79]. This process was based on the conversion of the substrate into lactones by prior grown cells of _W. lipofer_. After culture conditions optimization, this biotransformation process resulted in the maximum production of 51 g/L of \(\gamma\)-dodecalactone using 60 g/L of substrate after 40 h, with a yield of 85% and a productivity of 1.7 mg/L/h. These production parameters are by far superior to those obtained before, namely in patents [79]. Moreover, using a different substrate, namely a commercial 12-hydroxysteric acid, _W. lipofer_ was able to produce 27.6 g/L of \(\gamma\)-decalactone after 40 h [78]. According to the authors and to the extent of our knowledge, this is the process with the highest titers reported for a lactone produced by a microorganism. Another oleate hydratase from _Macrococcus caseolyticus_ that could be a good candidate for this type of process, namely for the conversion of oleic acid to 10-hydroxysteric acid, was also identified [80]. Following this trend towards more sustainable production of lactones, [PERSON] and colleagues exploited a new substrate. These authors tested the ability of _Y. lipolytica_ and _Lindurea saturus_ strains to produce \(\gamma\)-decalactone from crude glycerol [81]. Crude glycerol is the main by-product of the biodiesel industry and contains several contaminants, including fatty acids. For this reason, and despite not being discussed in the study, it seems that the lactones produced are derived from the fatty acids present in the substrate and not from the glycerol per se. There is no report that _Y. lipolytica_ can perform de novo biosynthesis of lactones [23], and it is equally unknown if _L. saturus_ can do it. Therefore, taking into account the high concentrations of crude glycerol used (100 g/L) and the maximum titer obtained in shake-flask fermentations (5.8 g/L by _L. saturus_) [81], one can assume that this production was based on the fatty acids present in this industrial waste. Nonetheless, crude glycerol is presented here as a more sustainable and available alternative for biotransformation. It is known that _Y. lipolytica_ is not able to produce lactones from oleic acid without genetic engineering [20; 23]. In turn, _A. gossyspii_ is able to biotransformation oleic acid and other non-hydroxy fatty acids into lactones. Characterization of the fermentation broths of industrial riboflavin (vitamin B\({}_{2}\)) production, where soybean oil is used as substrate, identified six different \(\gamma\)-lactones: three saturated, \(\gamma\)-nonalactone, \(\gamma\)-dodecalactone and \(\gamma\)-decalactone;three unsaturated, cis-6-\(\gamma\)-dodecenolactone, \(\gamma\)-jasmolactone and (\(Z\))-5-Decen-4-olide [10]. A total of 170 mg/L of total lactones were detected, cis-6-\(\gamma\)-dodecenolactone being the most predominant. Soybean oil is mainly constituted of linoleic acid, oleic acid, palmitic acid and linoleic acid, indicating that the fungus is able to biotransform unsaturated fatty acids into lactones. This reinforces the idea of [PERSON] and colleagues that _A. gossypii_ may be able to insert a hydroxy group into fatty acids.