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3.2 Enzymatic Hydrolysis and Box Behnken Model 3.2.1 Effect of Different Pretreatments on Enzymatic
Hydrolysis of Various Cellulose Substrates Plots in Fig. 4 showed that the glucose concentration
increased with reaction time. .It was also indicated that organosolv treated OPEFB cellulose produced
the highest glucose concentration followed by OPEFB treated with acetic acid, microcrystalline cellulose,
cellulose iber and OPEFB after deligniication.
3.2.2 Effect of Different Variables on Enzymatic Hydrolysis of Cellulose Extracted from OPEFB
A 15-run Box-Behnken design with tree factors and three levels, including three replicates at the centre
point, was used for itting a second-order responses surface. The three centre point runs were added to
provide as a measure of process stability and inherent variability. The considerable variation in the glucose
concentration from OPEFB cellulose under different conditions was shown in Table 2.
Figured 5 showed that model run experiment number 8 produced the highest amount of glucose,
that was 167.4 gL. In this 8 experiment, 0.2 ml enzyme and 0.7 g of samples has been used with 94
hours’ time reaction. The lowest glucose concentration was at run number 9 with concentration only 21.6
gL. The glucose concentration increased with the increasing amount of samples cellulose. The
longer the reaction time also increased the glucose concentration produced. The best total enzyme to
produce high glucose concentration was 0.2 ml. That means, used the same amount of novozyme 188 and
celluclast 0.1 mL: 0.1 mL could improve the activity of these two enzymes as catalyst. This is probably due
to the stability of enzyme activity. Figured 5 showed the effect of variable total enzymes a and the effect
of variable amount of samples b on the result of glucose concentration. Fig. 5 a showed that 0.2 mL
total enzyme produce highest concentration of glucose 167.4 gL and 0.1 mL produced the lowest glucose
concentration 21.6 gL. Fig. 5 b showed 0.7 gram of samples can produce the highest glucose concentration
167.4 gL and 0.3 gram of samples produced the lowest concentration of glucose 21.6 gL.
Table 2- Box-Behnken Design with Experimental and Predicted Values of Polysaccharide Yield
Run X1
X2 X3
Glucose concentration gL
1 -1
-1 91.8
2 -1
-1 97.2
3 +1
+1 97.2
4 +1
+1 93.6
5 -1
-1 55.8
6 +1
-1 39.6
7 -1
+1 122.4
8 +1
+1 167.4
9 -1
-1 21.6
10 +1
-1 97.2
11 -1
-1 48.6
12 +1
+1 100.8
13 52.2
14 54.0
15 55.8
Glucose released with the help of enzymes, celluclast and novozyme 188 were used as catalyst
in this hydrolysis reaction. Celluclast contained of Endoglucanase and Exoglucanase, enduglucanase
broke the cellulose in the middle of chains to reduce the degree of polymerisation and exoglucanase
worked at the ends of the chain to break the chain into cellobiose the dimer of glucose. Novozyme 188 is
cellobiase contained of β-glucosidase with functioned to break the cellobiose into glucose monomers [16].
Reation time h
10 20
30 40
50
G lu
co se
c on
ce nt
ra tio
n g
L
5 10
15 20
25 30
35 40
Cellulose fiber Cellulose microcrystalline
OPEFB after delignification OPEFB treated with Acetic Acid
Cellulose extracted from OPEFB
Fig.4. Effect of Different Pretreatments on Enzymatic Hydrolysis of Various Cellulose Substrates
ISBN : 978-602-17761-0-0
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4. Conclusion
