Team:Unesp Brazil/Hardware

iGEM Unesp Brazil



In the human body, the largest amount of microorganisms is found in the large intestine due to the favorable characteristics for their survival, such as the slow movement rate, pH and nutrients availability. The large intestine carries more than 101² microorganisms per body gram (Figure 1).

Figure 1. Schematic representation of the human gastrointestinal tract: the number of bacteria per gram of intestinal contents, nutrient availability and bacterial fermentative activity typically found in different sections of healthy individuals (Payne et al., 2012).

An important analysis to be carried out with genetically engineered probiotics is how they respond to the diversity of microorganisms present in the gut’s microbiome. For this analysis to be performed reliably and without the use of animals in initial stages of research, it was necessary to construct and model a bioreactor that resembles the human intestine and that has the ability to simulate, as closely as possible, the intestinal conditions. We designed our bioreactor to simulate the three parts of the large intestine, proximal, transverse and distal colon.

The widely used systems for bowel simulation are systems with a single reactor, continuous systems with one or more reactors, and artificial digestive systems (involving the stomach, small intestine, and large intestine). Batch fermentative systems are more difficult to control and are used for short-lived experiments, depending on the inoculum rate and rate of substrate consumption. Continuous systems are used for conducting long-term tests so that multi-stages are very interesting for the different parts of the intestine, which reaches the steady state becomes reproducible.

As a way of describing more efficiently the intestinal characteristics, a different reactor model was proposed, which will be divided into three parts: with piston flow as it occurs in the intestine, continuous mode of operation, sample withdrawal, control pH, temperature control system and also with proportional dimensions to real ones. To avoid washing the microorganisms, a porous support was used in the first part of the reactor.

3D Bioreactor Design and Assembly

AutoCAD 3D was used to draw the bioreactor for 3D printing (MakerBot Replicator +). The technique allows for greater versatility of forms and complete personalization of the new model. Designed and printed parts can then be assembled to create a fully functional bioreactor, minimizing problems related to the number of outlets, inputs, control and sampling points, connector sizes and leaks.

A photo of the first bioreactor designed, printed and assembled is shown in Figure 2. Its dimensions are presented in Table 1.

Table 1. Comparison of the dimensions of the human large intestine and the bioreactors system for the determined parameters.

Figure 2. Bioreactor 1.0.

We faced a few problems printing our bioreactor model 1.0, once its size (20 cm) is bigger than the printing area of our 3D printer. Therefore, we had to make changes in our project.

Our 3D printer builds objects layer by layer with melted plastic. Thus, complex structures are printed with supports to assist sustentation. Unfortunately, these supports affect the bioreactor functionality, lessen resolution, and cause leakage.

For the second attempt, we tried to make it simple and easy to print. Bioreactor model 2.0 was designed without the sample outputs and shorter (16 cm) (Figure 3). For this one, it is possible to see the designed fittings for sealing the reactor.

Figure 3. Bioreactor model 2.0.

As shown in Figure 4, we were able to print and assemble the model 2.0. However, once again it was leaky due to unforeseen supports.

Figure 4. Bioreactor model 2.0 printed and assembled.

After two failed attempts, bioreactor model 3.0 was designed and printed in several small pieces, which were glued and varnished afterwards. That way, we thought we could eliminate the supports and prevent leakage. However, the structure built by the 3D printer was loose and allowed water to permeate the mesh.

Figure 5. Bioreactor model 3.0.

At that point, we decided to change completely our approach. We combined silicone hoses with 3D printed pieces to build the Bioreactor model 4.0 (Figure 6).
Finally and fortunately, our Bioreactor model 4.0 has no leakage and it is fully functional!

Figure 6. Animation showing the parts and the assembly of bioreactor model 4.0.

Figure 7. Bioreactor 4.0 printed and assembled.

Final test

A 12h test was performed to evaluate bioreactor 4.0 impermeability and resistance to 37°C, which is the temperature of the human body. A mixed culture was grown in the bioreactor operated as a chemostat with continuous in- and outtake. The bioreactor was also stable throughout the time, supporting bacteria growth. We were even able to raise the temperature up to 60°C without causing any structural damage.

Our bioreactor model 4.0 costed about USD 40. Cost was estimated in Reais (Brazilian currency) and converted to US Dollars. The price of PLA and the silicone hoses may be different in other countries.

Figure 8. Time to test them!

Table 2. Cost of the materials used to build the bioreactor in US$.

Click here to download the files and build your own bioreactor!


School of Pharmaceutical Sciences | Chemistry Institute


School of Pharmaceutical Sciences
São Paulo State University (UNESP)
Rodovia Araraquara Jaú, Km 01 - s/n
Campos Ville
Araraquara, São Paulo, Brazil