Water Filtration

Overview

By attaching functional proteins to cellulose we can expand it's properties and selectivity capture specific contaminants in water. We used five different cellulose binding domains and fused them to different metal binding proteins, and sfGFP. We performed assays to test the binding of the CBD fusions to our cellulose.

Key Achievements

Made cellulose binding domains

Introduction

Water is typically purified by passing it through layers of porous materials, each specially selected for its ability to remove specific forms of contamination. All porous materials can filter particles by size, many have extra capabilities thanks to their chemical properties. For example, charcoal - or activated carbon - is a porous component of household and industrial water filters that can also bind large or electron-rich molecules via the van der Waals forces, and catalyse the breakdown of other chemicals such as molecular chlorine. While there are many different types of filters, we can categorise and compare them using their measurable physical and chemical properties. The key physical properties are:

Pore size - average or maximum size of pores in the material
Porosity - volume of the filter not occupied by solid material
Tortuosity - length of paths through the filter compared with a straight line
Adhesion - the strength of hydrogen bond interactions between the fluid and filter
Kinetic rate constants - parameters defining how the filter material affects chemical reactions in the fluid

These properties contribute to the filter’s flow rate, measured in units of fluid filtered per unit surface area of the filter per unit time (e.g. Ga m^-2 h^-1). Altering the filter’s flow rate generally affects the filter’s efficiency at removing its targets, as increasing the flow rate typically involves increasing the pore size or reducing the surface area of the filter in contact with the fluid. This trade-off is the main hurdle in industrial water filtration, where demands for high-quality filtration require very low flow rates. Industrial processes are further complicated by blockage of the pores, fouling of the filter by organisms and the lack of a complete set of materials that will exhaustively filter all contaminants - heavy metal ions, small organic molecules and non-polar compounds remain difficult to filter without prior chemical treatments that themselves need to be filtered out.

Turbidity

Ultrafiltration membranes remove particulates by size exclusion. Our bacterial cellulose membranes completely removed turbidity from input water outputting clean, potable water. Input water containing 1% w/v organic solids from Imperial College Queens lawn was vortexed then 3ml passed through filters of 25cm3 area. Turbidity of flowthrough was measured in a spectrophotometer, absorbance at OD₅₀₀ as recommended by Epa (1993).

Figure 1: The picture shows different amount of colouring in the water representing the debris left after filtering water mixed with dirt using different filter methods. From left to right: Unfiltered water, tissue paper filtered water, filter paper filtered water, bacterial cellulose filtered water, clean distilled water.

Cellulose sheets produced by G. xylinus iGEM were dried to thin filters around 20 g/m³. Coffee press filtration with these filters was compared to similar filtration by tissue paper and VWR qualitative medium filter paper (5-13 um pore size).

Average OD₅₀₀ values from three replicates:

Water unfiltered: 0.072.
Tissue Paper filtered: 0.039.
Filter paper filtered: 0.023.
Bacterial Cellulose filtered: 0.00.
Distilled water: 0.00.

Cellulose functions as a water filter and completely removes suspended particles.

Phytochelatin-dCBD metal binding assay

Figure 2: This table summarises the metal concentration in EDTA after washing cellulose samples exposed to different treatments: different phytochelatin-CBD fusion protein lysates added and the salt solution it was incubated in. Concentrations were obtained via Inductively coupled plasma optical emission spectrometry (ICP-OES). The lower part of the table shows secondary treatment of the cellulose after the first EDTA wash.

Figure 3: Graph to compare the different cellulose treatments and their correspondence to the average metal concentrations after the first EDTA wash.

Figure 4: Graph to compare the different cellulose treatments and their correspondence to the average metal concentrations after the second EDTA wash.

Figure 5: This graph compares the averages between the first and second wash for each sample type, showing that all but one of the samples reduced in metal concentration in the second wash.

Nickel filtration assay

The nickel ions are an example of heavy metals, poisonous even in relatively small concentrations in water and notoriously difficult to filter with current filtration methods. Therefore our filtration concept was tested against a concentration of nickel in water that far exceeds the safe limits. We have attempted to filter high amount of nickel (250 μM) through the cellulose filters grown by the G. Xylinus ATCC53582 strain (K1321305). The phytochelatin-dCBD fusion (K1321110) was coated on the surface of the cellulose to make a nickel specific functionalised ultra filtration membrane. To test the two membranes we have used coffee press. As a control measure we have also attempted to the filter the nickel solution through the cellulose that was not further functionalised. Obtained filtrates have been quantified and analysed using Mass Spectrometry.

Results

Figure 7: Here shown is the decrease in the nickel content of the filtered nickel solution. The filtrate was obtained afer filtering with the phytochelatin-dCBD functionalised cellulose membrane. The concentration of unfiltered solution is provided as control.

Figure 8: Here shown is the difference in the decrease of the nickel content when filtered with cellulose compared with the functionalised cellulose with phytochelatin-dCBD.

From the results in figure 7, it can be seen that the cellulose functionalised with phytochelatin-dCBD (K1321110) is able to decrease the nickel content in the filtrate by 95%. This shows that the ultrafiltration method with functional protein domains on the cellulose is working and is a viable option for heavy metal removal from polluted water. Also the results observed in figure 8 shows that the non-functionalised cellulose is able to remove nickel from water, and the functionalisation of the cellulose further increases the incremental decrease of the nickel content in the filtrate by an approximate further half. The results presented suggest that bacterial cellulose is a good method for removing nickel from water and functionalisation of the cellulose provides a further decrease in the final concentration of the nickel filtrate.

This mechanism shows that the cellulose itself is able to remove contaminants from water, in this particular example Ni²⁺. It shows good ability to remove nickel ions, possibly due to nickel ion small size and hydrophillic nature. It is also shown that the functionalisation of the cellulose with the specific heavy metal binding chelating protein fusion with a cellulose binding domain provides an extra incremental decrease in the Ni²⁺ concentration found in the filtrate. It is possible that depending on the nature of the contaminant, the incremental change provided by the functionalisation step can be increased. This can be of specific importance for the contaminants that cannot be caught by the inherent filtration ability of the cellulose material.

Team:Imperial/Water Filtration

From 2014.igem.org

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