## 5 Liquid Mycelial Cultivation of Macrofungi: Methods and Strategies  
Global production of mushrooms grew from 6.90 to 10.24 million metric tons from 2008 to 2017 [168]. There is a great range of nutraceutical, pharmaceutical, and novel food products containing macrofungi fruiting bodies or mycelium in Asia and the USA and a growing market in the rest of the world [118]. Solid-state cultivation of fruiting bodies requires several months, a large volume of substrate and space, and the quality of the final product can be difficult to control [169]. In contrast, the submerged culture of macrofungi promises a faster, more efficient, and more consistent alternative for the production of fungal bioactive metabolites [88]. Submerged cultivation is any strictly controlled and monitored biochemical process where a liquid nutrient substrate is enzymatically converted by microbial species [60]. Submerged cultivation in bioreactors is considered the best method for producing bioactive molecules with medicinal properties from macrofungi. Process monitoring, control, reproducibility, and the high product quality required for such production can only be achieved in bioreactors [75; 170]. That is because submerged liquid cultivation allows for the use of analytical tools that can supply at-line and online data, and thus a better characterization and modeling of the bioprocesses has been achieved in the last decades [171; 172]. [PERSON] et al. [173] performed the first reported submerged culations of macrofungi in 1966, where they tested hundreds of Basidiomycota in search of antitumor substances. Table 2 presents the content of different bioactive metabolites in the fruiting bodies of the mushroom compared to that of the mycelium of the same species. Several studies indicate that the cell wall composition of mycelium can be similar to that of the mushroom fruiting body [174], whereas [PERSON] et al. [118] show that in many cases, the mycelium contains more bioactive compounds than the carposome, e.g., carbohydrates in _P. ostreatus_ (70.4 vs. 27.7%), lipids in _L. edodes_ (20.0 vs. 4.0%), or sterols in _Tuber sirense_ (5740.5 vs. 1883.0 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro g}\)). In addition, a study comparing the chemical composition of the fruiting bodies with the submerged cultured mycelium of _C. militaris_ demonstrated that the mycelium had a higher content of polysaccharides, phosphorus, cordycepin, cordycepic acid, alanine, and vitamins B3 and B2 [175]. There are more studies that confirm that macrofungi can produce extraordinarily high amounts of EPS in a small period of time under submerged cultivation conditions [94]. The mycelium biomass of _P. ostreatus_ produced in a batch stirred-tank bioreactor contained enhanced quantities of bioactive metabolites known to be found in fruiting bodies, thus proving that submerged cultivation can be a promising alternative [176]. Advantages of liquid mycelial cultivation of macrofungi [177, 178, 179]. A wide range of products from a wide range of fungi can be produced, while many products are produced best in submerged conditions. Additionally, there are the following advantages:  
* Kinetic, heat, and mass transfer parameters can be estimated;
* Easy control and adjustment of the operating parameters;
* Very good reproducibility when the medium composition is characterized;
* Defined medium allows for easier purification of desired substances;