To investigate the sugar concentration of different types of biomass, and thus determine their usability as biomass sources of sugar.
Biomass products with characteristics which suggest high sugar content such as a sweet taste, aroma or sticky texture would have higher sugar content than the other products, with less evident characteristics. Thus, fruits are predicted to have a higher sugar concentration than grain and grass biomass types.
A 3,5-dinitrosalicylic acid (DNS) assay was conducted to estimate the total amount of reducing sugars derived from pretreatment of lignocellulosic biomass sources. The samples tested were compared to a glucose standard curve. The samples tested included
Starchy waste: Bread
Mixed: Peach; Banana; Coconut Pulp; Coconut Water; Strawberry
Fibrous: Sawdust; Barley Straw; Bamboo Bark; Sifton Bush; Wattle; Wheat Grain; Barley Grain; Sorghum Grain; Agave
Results:
In this sugar extraction experiment, we are investigating the concentration of reducing sugars in different biomass sources. The sources which we have selected are entries in the biomass waste encyclopaedia, which were assessed to be highly suitable as sugar sources. These biomass were ones which were researched to be commonly wasted - domestically and industrially, have a high sugar content, and have high availability in the market. Through this practical procedure, we aimed to determine the concentration of reducing sugars in these sources and verify their suitability for use as biomass sugar sources. The biomasses which are ultimately most productive and suitable will be highly versatile and valuable sources of biofuel, serving the purpose of our project which is to reduce waste while enhancing the efficiency of sugar intake in bacteria.
DNS assay is a method which is widely used to determine the concentration of reducing sugar. DNS, abbreviated for 3-5 dinitrosalicylic acid, is an aromatic compound which is used to treat reducing sugars. This assay tests for the presence of free carbonyl group (C=O), the so-called reducing sugars.This involves the oxidation of the aldehyde functional group (in glucose) and the ketone functional group (in fructose). When they react in boiling water, the 3-5 dinitrosalicylic acid is converted to 3-amino-5-nitrosalicylic acid. This chemical strongly absorbs light at 540 nm, which gives it a brown-orange colour. By measuring the intensity of the orange colour - optical density, it can be used to compare the concentration of reducing sugars in different samples. As the concentration increases, the intensity of the colour will also increase. Thus, the DNS assay effectively allows for the investigation of the reducing sugar concentration in different types of biomass.
The results of this sugar extraction analysis procedure are pivotal in determining the right and most cost effective and efficient resource to use for sugar extraction, where efficient bio-waste may prove to be too expensive to convert into energy, cheap bio-waste could have a low sugar content overall making the whole process futile. Although we initially expected fruits to contain the highest amount of sugar, after the DNA assay it was discovered that the sample with the highest sugar content turned out to be wattle, followed by bananas and bread, this result was partially expected apart from the wattle. When handling the wattle sample, the wattle presented a dry and almost dead consistency, similar to that of dry leaf or branch, however it seemed that the internal composition of the wattle contained high amounts of sugar. Post sampling research revealed that seeds within wattle actually release sugar over time which would attract bees and other insects, this process of the slow release of sugars could have been expedited through the sugar extraction process thereby producing the highest score for the sugar levels.
Second highest value being the banana was hypothesised. Where due to fruits' sweet taste and aroma it was natural for bananas to contain a high concentration of sugars. One factor that would have affected this was the texture of the banana, where its soft flesh of the banana could’ve been effectively broken up evenly for maximum surface area contact with the acid, where other samples such as the coconut pulp had a highly hard texture that was difficult to break into sufficiently small pieces to maximise surface area.
The third highest value of bread was also predicted since the bread used was standard commercial bread which contains sugar in production. Within the food industry sugars are added into nurmours products (including bread) that supports the combination of extended shelf life, adding texture and flavour and assisting with browning. Like salts, sugar also acts as a preservative against mould and also helps bread keep its moisture.
This shows that common household food waste would benefit greatly from our gene. Intriguingly, the high result for wattle allows us to consider applications in incentivising the public to collect fallen parts of the largely ornamental plants, which often become a fire hazard in Australia.
It can be concluded that the best sources are wattle, followed by banana and bread respectively. These three biomass sources are the most suitable options for biofuel to be used by bacteria, as they have the highest reducing sugar concentration. As these three biomass sources are also ones which are commonly wasted and have high availability in the market, they serve as promising candidates for bacteria usage. By employing these biomass sources, we can effectively reduce wastage of these sources around the world, while providing valuable fuel sources for bacteria, to be used for a variety of bio-industrial uses.
Our experiment aims to improve the efficiency of sugar intake in bacteria, by allowing for the consumption and absorption of more types of sugars, which are commonly found in natural biomass sources. For our experiment, we are investigating Escherichia coli. Our synthetic biology modifications have successfully increased the efficiency of sugar intake under lab conditions. This experiment has highlighted the most potent natural biomass sources. Naturally, the next step is to investigate the improvement of E. coli, using natural biomass sources. For example, further testing could involve feeding wattle - the best biomass from this experiment to improved E. coli and non-improved E. coli, thus determining the applicability of our genetic modifications in the real world, with natural biomass sources.
Using the trends found in this experiment, further biomass sources with high potential can be found and investigated. For example, looking for biological characteristics closely related to the biomass sources which have already been proven to contain high concentrations of reducing sugars could uncover new biomass sources, which can be used for their sugar concentration.
Another way to further the experiment is to conduct a more advanced analysis of the reducing sugar concentration. This can be done through Gas Chromatography Mass Spectrometry. The equipment is currently not available to our team, but it could potentially be an opportunity to collaborate with other teams.
Reliability refers to the consistency between repetitions of an experiment. Overall it can be concluded that this experiment is considered reliable. The method of testing is detailed and highly repeatable, so that external reliability can be tested to verify data.
Validity refers to whether variables were properly controlled within an experiment so that a relationship could be established between the independent and dependent variables. The independent variable was the biomass used, and the dependent variable was the quantity of its reducing sugars by weight. Controlled variables such as the weight of each sample and volume of acid used in testing was kept consistent.
Accuracy refers to how close the acquired measurements were to the actual value. Since a professional scientific scale was used down to the microgram and calibrated lab measuring equipment was also used we can conclude that the experiment was accurate. Higher levels of accuracy can also be achieved in a higher level of laboratory setting.
Through this experiment, the sugar concentration of different types of biomass was successfully investigated. Thus, the suitability of different biomass sources was determined and the options for the best biomass sources was further narrowed down. It is concluded that wattle has the highest reducing sugar concentration, followed by banana and bread. Biomass with the least reducing sugar concentration included coconut pulp, strawberry and sourghan. The hypothesis was proven incorrect, as there wasn’t a strong, direct correlation between the presence of sugary characteristics (taste, smell, texture) and the actual concentration of reducing sugars. Rather, grains and grass biomass types were highly underestimated, and have been concluded to be highly potent biomass sources.
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