the hydrolysis of cellulose

Cellulose hydrolysis is also an important factor in biofuel development because bacteria absorb glucose rather than cellulose during fermentation to produce biofuel. Because of the existence of crystalline compounds, the hydrolysis of celluloses is complicated. Because of the thick packing of cellulose chains, these structures behave in this manner and are very stable under a variety of chemical conditions. Cellulose can only be chemically and enzymatically hydrolyzed.
The enzymatic approach involves the use of bacteria-secreted proteins to hydrolyze cellulose, which involves a large number of enzymes – cellulose. Research has shown that these enzymes play different roles cooperatively in the hydrolysis of cellulose: some cleave the cellulose chain from the middle into fragments containing 4-5 glucoses, some breakdown these fragments into smaller units of two glucoses, and some finally turn these small units into single glucose. (Hames, 2009).

Hypothesis:

In this lab, cellular hydrolysis experiment was carried out for different concentration, conductance, diafiltration, cross filtration, BSA filtration rate of absorbance until dynamic equilibrium was reached and all the aspects and factors have been explored exhaustively.

Materials:

Exercise 1A:

The materials used include a 30cm piece of 2.5cm dialysis tubing, string, scissors, 15mL of 15% glucose/1% starch solution, 250mL beaker, distilled water, and 4mL of Lugol’s solution (Iodine Potassium-Iodine or IKI).

Exercise 1B:

This exercise required six 30cm strips of presoaked dialysis tuning, six 250mL cups or beakers, string, scissors, a balance, and 25mL of  these solutions: distilled water, 0.2M sucrose, 0.4M sucrose, 0.6M sucrose, 0.8M sucrose, and 1.0M sucrose.

 Exercise 1C:

The materials that were required include 100mL of these solutions: distilled water, 0.2M sucrose, 0.4M sucrose, 0.6M sucrose, 0.8M sucrose, and 1.0M sucrose, six 250mL beakers or cups, a potato, a cork borer, a balance, paper towel, and plastic wrap.

 Exercise 1D:

 The materials used include a calculator, and a pencil.

Procedure:

Exercise 1A:

In this experiment, the concentration was lowered by use of distilled water while the conductivity was being monitored and recorded after every dilution of the solution. It started from 100% high concentration to 60% while each conductivity was being measured against particular given concentration.

 Exercise 1B:

In this section, aqueous buffer was used to obtain flux rate data under different conditions. 2 flat‐bed cross flow filters was used, with different MW cutoffs. The final results were then taken under proteinaceous solution so as to simulate an actual bioprocess solution.

Exercise 1C:

The experiment in this section is about the absorption of glucose. The information of graph and the tabulations are in figure 3.

Exercise 1D:

This experiment as shown in figure 4 was done to determine the activity of cellulose on cellocast and the resulting table for both activities were drawn

Results:

Exercise 1A:

Figure 1 Graph of conductivity against concentration

The above diagram was drawn from the results of concentration versus conductivity using filtrate collected in ml against permeated conductivity. It is a result of determining the extent of buffer exchange using the conductivity data. The assumption here is that the solution remained contaminated free throughout the experiment and there is no faults in the apparatus used to measure conductivity.

  Exercise 1B:

Table 1 Results obtained from Diafiltration experiment

The pressure of activities caused by enzymes activities were tabulated during the experiment

Table 2 data obtained from 0.5^2 cross filtration

Figure 2 figure showing both tabulation of data and a graph showing the results obtained from the activity.

This section of the experiment was conducted to determine the rate of absorption of glucose and a graph was drawn and therefore it is good to say that the rate of absorbance is directly proportionally to the amount of glucose.

Figure 3 Figure showing both the table of cellobiase and cellocast absorbance and consequent graph drawn against time for both.

General

Experiments were conducted and data to monitor the concentrating effect of the filtration operation were collected. A run condition was set up and was operational over time, collecting filtrate, estimating flowrate and monitoring optical density at 280 nm to estimate protein concentration in the final run where Protein is used and in the diafiltration run with protein.

For the diafiltration, simulation of buffer exchange from a high‐salt buffer to a low salt buffer was carried out typically to desalt elements from ion‐exchange chromatography

Finally, an experiment was carried out to determine the activity of a Cellobiase on Celluclast. This section shows how enzymes breaks a 1,4 glucosidic linkage between 2 glucose monomers in the final step of cellulose hydrolysis. Glucose results in experiment 1 was used to estimate the cellobiase on Celluclast and the glucose yield for each reaction.

Conclusion:

Cross‐flow filtration is an important bioprocess for the partitioning of process streams. Microfiltration is used to remove or concentrate particulates, including cells and cell debris. Ultrafiltration can be used to concentrate high molecular weight components or remove high molecular weight contaminants from smaller product molecules (Deguchi, Tsujii & Horikoshi, 2006). Finally, these devices can be used to facilitate buffer exchange prior to chromatography, in a process known as diafiltration.

Cellulose hydrolysis is key to second generation ethanol processes, whereby fermentable glucose is developed from a combination of thermochemical and enzyme treatment of lignocellulosic materials. Lignocellulose comprises of cellulose, hemicellulose and lignin. The cellulose and to some extent the hemicellulose fraction can be hydrolysed to release fermentable glucose by cellulases. (Deguchi, Tsujii & Horikoshi, 2006)

References

B. R. Hames: “Biomass Compositional Analysis for Energy Applications,” Methods Mol. Biol. 581, 145 (2009).

S. Deguchi, K. Tsujii and K. Horikoshi. “Cooking Cellulose in Hot and Compressed Water,” Chem. Commun. 61, 3293 (2006).