Wednesday, 16 September 2009

Microfluidic System Engineering for Enzyme Analysis in Genetically Modified Sugarcane

Schematic of the components of Integrated Cellulase Unit, ICU

Motivation and Drivers

Increasing and continuing dependence on dwindling petroleum resources has lead to an increase in oil prices and the concentration of green house gases. Thus, it is desirable to make a transition from non-renewable resources to renewable resources such as biofuels (e.g. bioethanol, biodiesel), solar, wind, and hydro.

Bioethanol is believed to have a potential to decrease our dependence on petroleum-derived transportation fuels. Currently, sugarcane in Brazil and corn in the United States are the most commonly used feedstocks for the commercial production of ethanol (first generation biofuels). But their use has been discouraged due to fears that the use of food crops (such as corn and sugarcane) for ethanol production can increase food prices and decrease food supply. However, the conversion of lignocellulosic biomass (second generation biofuels) to ethanol does not create a pressure on existing food or land resources, and therefore presents an ideal opportunity for lucrative commercial bioethanol production.

One source of lignocellulosic biomass is bagasse (the fibrous portion left after stalks of sugarcane are crushed to extract the juice). It has been estimated that with just 50% of the bagasse that is currently being produced from the sugarcane crop in Australia approximately 1.2 billion litres of ethanol can be produced per year. This is equivalent to ~2% of the country’s gasoline consumption in the years 2006.

Hydrolysis of cellulose and hemicellulose in pretreated bagasse requires a number of cellulase and hemicellulase enzymes. The cost of these enzymes is ~30-50 US cents per gallon (~ 10 US cents/ litre) of ethanol and has been as the key barrier to economic production of ethanol from bagasse. Thus, it is necessary to reduce the cost of enzymes for the production of ethanol from bagasse to be economically viable.

It has been suggested that genetically modified (GM) plants which can express multiple cellulases and hemicellulases have the potential to provide lower cost performance cellulosic enzymes. But the concentration of enzymes in different batches of sugarcane is expected to vary with harvesting and cultivation conditions, thereby necessitating the characterization of every batch of cane juice for: 1) Fiscal measures and 2) Process control.

Thus, the aim of this research project is to develop an instrument (called Integrated Cellulase Unit, ICU shown in the figure above) to enable real-time analysis and monitoring of the concentration of multiple cellulases in every incoming batch of GM sugarcane.

ICU Analyser

In a search for an appropriate basic principle for the ICU analyser, a review of the capabilities of High Performance Liquid Chromatography (HPLC), Capillary Zone Electrophoresis (CZE), Isotachophoresis (ITP) and Isoelectric focussing (IEF) have been evaluated.

At this stage of the project, ITP is the choice of technique for the ICU analyser due to its several advantages which includes its ability to concentrate dilute samples, higher sample loading capacity, lower susceptibility to blockages, relaxed requirements on sampling and detection units, and possibility for automation. These advantages are of utmost importance for monitoring the concentration of enzymes in a process environment.

Future work will focus on lowering the limit of detection of these enzymes.

Research Partners and Principal Contacts

Ms Ruchi Gupta, Syngenta Sensors University Innovation Centre, University of Manchester, UK

Prof Peter Fielden, School of Chemical Engineering & Analytical Science, University of Manchester, UK

Prof Nick Goddard, School of Chemical Engineering & Analytical Science, University of Manchester, UK

Mr Ian O'Hara, Centre for Tropical Crops and Biocommodities, QUT, Australia

Would you like to become a research partner? Are there other aspects of this research that we should be bringing in?