Team:NCTU Formosa/Wet Lab/Curcumin Biosensor

After choosing α_S1-Casein as our biosensor, we should choose the method to detect curcumin. We choose the electrochemical impedance spectroscopy (EIS) and Differential Pulse Voltammetry (DPV), the two detecting methods in electrochemistry.

EIS is simple, convenient, and rapid, so that it is quite suitable for detecting whether the biosensor effective or not. This method uses the principle that impedance will be changed by charge transfer, and then measure the change of impedance by Alternate Current. Therefore, if we find out that our measured result has the stronger change than negative control, we can say our method could produce a fierce Redox reaction. However, because EIS is not a suitable method for detecting small molecule, the error will be very large if we choose it to perform formula computing. So we decide to use DPV to improve this problem.

DPV uses the difference between before and after the pulse application in order to solve the influence of background noise. This principle is difference from EIS. We hope we can find out which method is more sensitive to curcumin, and create more accurate formula.

Data Analysis

First of all, we use EIS to check whether our biosensor can detect curcumin. We have two samples. Red line is connected to general chip, and blue line is the chip connected with α_S1-Casein(Fig 1). When our biosensor is connected with α_S1-Casein to detect curcumin, impedance value become larger. That is our biosensor connected with α_S1-Casein produce more fierce Redox reaction than another. This figure also represent that the biosensor connected with α_S1-Casein have more effect of detecting curcumin than none.

Figure 1: The method EIS has poor sensitivity for curcumin detection.

After we prove that adding α_S1-Casein can detect curcumin, we hope we can use more accurate method to show our data. We choose DPV to replicate the experiment above, and determine which method is sensitive. Comparing with fig1 and fig2, we can find out that chips with α_S1-Caseinis almost no change in EIS experiment, and change a lot in DPV experiment in contrast. In this way, we can explain that DPV is more sensitive and has better signal than EIS.

Figure 2: The sensitivity test of curcumin Biosensor in Dpv.

Doing same experiment by using two different kinds of methods, we know that DPV is more sensitive. Then, we want to know what condition is more suitable to detect curcumin. We use PB and PBS, two buffers, as curcumin solvent. In fig3, blue line is use PB Buffer, and red line is use PBS Buffer. In the result, LOD (limit of detection) of curcumin in PB buffer is 10pM and in PBS buffer is 1nM. This means that the sensitivity by using PB buffer is one hundred times than using PBS buffer. We also can confirm that NaCl will increase noise in our method. In the conclusion, we use the PB buffer as our solvent rather than the PBS buffer.

Figure 3: The improvement test of curcumin Biosensor in Dpv.

Figure 4: Curcumin concentration take natural logarithm to acquire polynomial.

Now, we know that connecting our biosensor with α_S1-Casein and using PB buffer are the best condition to detect curcumin by DPV. We use the data in this condition to curve fit. Fig4 easily shows that it is the Logarithmic function so we put the curcumin concentration into the Natural logarithm, and do the polynomial curve fitting. We can obtain the result in Figure 5, R² =0.9995. This represents our curve is really close to real value and get the following formula.

Figure 5: The detection result of real samples from turmeric.

We use our biosensor and estimated formula to detect curcumin. We mill the turmeric and divide into two groups to detect. One adds water and isn’t extracted, and another adds water and is extracted. The sample which isn’t extracted cannot be detected. It is true because curcumin exist in cell. Actually, this result also represent our sensor have strong specificity. That is our sensor will not be disturbed even if taking the whole turmeric to detect.

Conclusion

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