## 4 Discussion
This inconsistency is due to the protein assessment method. In the crude protein assessment, the total nitrogen content of the product is measured by Kjeldahl, and protein is calculated based on nitrogen content [33]. Therefore, any nitrogen from non-protein sources results in overestimating the protein content [33]. However, calculating the protein content by summing up the amino acids (resulting from protein hydrolysis) does not have this limitation [33]. It is worth mentioning that the crude protein content of wheat flour varies between 10.5% to 13.5% [34]. Thus, the protein content of oyster mushroom cultivation residue is not less than wheat flour. According to [PERSON], [PERSON] and [PERSON] [10], the total amino acid content of the oyster mushroom (grown in BSG and wheat bran mixture) was 34.8 g/100 g, and its essential amino acid content was 12.7 g/100 g. The aspartic acid, glutamic acid, alanine, valine, leucine, lysine and arginine content of the oyster mushroom was more than 2 g/100 g, and its glutamic acid content (5.3 g/100 g) was higher than other amino acids [10]. In fact, glutamic acid is the most abundant amino acid in oyster mushroom [35]. This can justify the decrease in the glutamic acid content of the cultivation residue. The comparison of amino acids between the initial substrate and cultivation residue indicated a decrease in most amino acids (except for arginine and threonine), particularly glutamic acid. It is worth mentioning that when the mushroom grows, it uses the available protein in the substrate and also produces its own protein. Therefore, the change is due to the different amino acids composition in the substrate and the mushroom proteins. In terms of essential amino acids in cultivation residue, although some of them were slightly higher (e.g., lysine, threonine) or lower (e.g., leucine) than wheat flour, the total essential amino acids content of this residue is similar to wheat flour [36]. Regarding the vitamin content of the cultivation residue, the niacin content of the substrate noticeably increased after oyster mushroom cultivation. Since the oyster mushroom has high niacin (90 mg/100 g) content [10], it can be presumed that the substrate is naturally fortified with niacin after cultivating oyster mushrooms due to the presence of mycelium. Although niacin has several health benefits [37], its upper limit of intake is 35 mg per day, and excess intake can result in vasodilatory effects (flushing) [38]. Since 100 g of oyster mushroom cultivation residue contains 42 mg of niacin (above the daily upper limit), this quantity of cultivation residue should not be consumed in a day. The niacin content of wheat flour is 1.6 mg/100 g [34]; thus, developing a bread prototype from a mixture of wheat flour and cultivation residue flour reduces the niacin content of this bread to the safe level. The cultivation residue's thiamine, riboflavin and pyridoxine content have slightly changed compared to the initial substrate; however, the quantity of these vitamins is not noticeably different from wheat flour [34]. Regarding lutein and zeaxanthin, although wheat bran and barley malt contain lutein and zeaxanthin [39; 40], the quantity of these nutrients was negligible in the cultivation residue. Since lutein and zeaxanthin were observed in brewery trub [41] and were not detected in cultivation residue, it can be presumed that they have been lost during the beer production process. The total dietary fibre and beta-glucan content of cultivation residue were higher than wheat flour. The cultivation residue's dietary fibre was 59%, whereas wheat flour's dietary fibre is 3% [34]. Therefore, the fibre content of cultivation residue is higher than wheat flour, and this residue can be considered a good source of dietary fibre. In terms of beta-glucan, the cultivation residue's beta-glucan was more than 6%, whereas wheat beta-glucan is less than 0.7% [42]. Thus, the beta-glucan content of cultivation residue is noticeably higher than wheat flour.