## 3 Results and Discussion
### Optimization of Fermentation Processes  
#### 3.2.1 RSM Optimization of Dilute Acid Pretreatment  
The central composite design (CCD) model with four factors for optimization of dilute acid pretreatment was used to create a regression equation (Equation S2a) as a prediction for maximum monosaccharide yield after input of values for response variables. The significance of factors and the interactions between factors on the composition of the hemicellulose hydrolysate sugars are shown in Figure S3a. Surface plots of interactions between factors on monosaccharide yield and production of inhibitors are shown in Figure S3b.  
The optimal parameters for efficient dilute acid pretreatment include a treatment time of 45 min, temperature of 125 \({}^{\circ}\)C, use of 1% (_v/v_) sulfuric acid, and 6.44% (_m/v_) biomass loading. Using these parameters for dilute acid pretreatment of vetiver grass, it is predicted that 20.07 g of total sugars per liter can be achieved for the hemicellulose hydrolysate. Upon pretreatment of vetiver biomass (n = 3), the hemicellulose hydrolysate was found to have 41.085 \(\pm\) 0.046 g of total sugars per liter, a 204.7% percent yield of the predicted. Inhibitors were produced above the targeted values at 0.305 \(\pm\) 0.031 g/L furfural, 0.197 \(\pm\) 0.035 g/L 5-hydroxymethylfurfural, and 4.859 \(\pm\) 0.070 g/L acetic acid.  
Optimization of dilute acid pretreatment of vetiver grass using RSM proved that acid concentration was the only evaluated treatment factor that had a negative impact on monosaccharide recovery from hemicellulose. Acid concentration also had the greatest effect on furfural production of the four tested treatment factors. The optimum sulfuric acid concentration of 1.0% (_v/v_) was most dilute out of the acid concentrations evaluated. Higher acid concentrations have been shown to decrease xylan hydrolysis, which results in lower xylose recovery during pretreatment [56]. However, inhibitor (furfural and acetic acid) yields have been found to increase with increasing acid concentration for dilute acid pretreatment [55; 56; 57]. Decreases in concentrations of 5-hydroxymethylfurfural were observed as acid concentration increased. Acidic conditions cause 5-hydroxymethylfurfural to degrade to levulinic acid and formic acid [58].  
Temperature was also found to greatly impact furfural production. Furfural is a degradation product of xylose which constitutes the majority of hemicellulose as the polymer xylan [56]. The toxicity of furfural was observed in Saccharomyces cerevisiae, specifically through the inhibition of dehydrogenases, which negatively impacts glycolysis [59]. Higher treatment temperature decreased the presence of 5-hydroxymethylfurfural, most likely due to further degradation to levulinic acid and/or formic acid [58]. Acetic acid formation was not significantly impacted by changes in temperature. Increases in treatment temperature have also been shown to decrease reducing sugar yields [56]. The optimal temperature for maximum reducing sugar recovery and production of below target concentrations of inhibitors for the delignification of vetiver biomass was 125 \({}^{\circ}\)C.