Dear Deepak,

In a recent study  polyaniline (PANI) film was successfully employed in a simple visual ethanol biosensor based on alcohol oxidase:

A Simple Visual Ethanol Biosensor Based on Alcohol Oxidase Immobilized onto Polyaniline Film for Halal Verification of Fermented Beverage Samples.

Bambang Kuswandi, Titi Irmawati, Moch Amrun Hidayat, Jayus  and Musa Ahmad

Sensors 2014, 14, 2135-2149; doi:10.3390/s140202135

Abstract: A simple visual ethanol biosensor based on alcohol oxidase (AOX) immobilised onto polyaniline (PANI) film for halal verification of fermented beverage samples is described. This biosensor responds to ethanol via a colour change from green to blue, due to the enzymatic reaction of ethanol that produces acetaldehyde and hydrogen peroxide, when the latter oxidizes the PANI film. The procedure to obtain this biosensor consists of the immobilization of AOX onto PANI film by adsorption. For the immobilisation, an AOX solution is deposited on the PANI film and left at room temperature until dried (30 min). The biosensor was constructed as a dip stick for visual and simple use. The colour changes of the films have been scanned and analysed using image analysis software (i.e., ImageJ) to study the characteristics of the biosensor’s response toward ethanol. The biosensor has a linear response in an ethanol concentration range of 0.01%–0.8%, with a

correlation coefficient (r) of 0.996. The limit detection of the biosensor was 0.001%, with reproducibility (RSD) of 1.6% and a life time up to seven weeks when stored at 4 °C. The biosensor provides accurate results for ethanol determination in fermented drinks and was in good agreement with the standard method (gas chromatography) results. Thus, the biosensor could be used as a simple visual method for ethanol determination in fermented beverage samples that can be useful for Muslim community for halal verification.

To view the methods section, please use the following link:

https://encrypted.google.com/url?sa=t&rct=j&q=&esrc=s&source=web&cd=2&cad=rja&uact=8&ved=0ahUKEwiT-dqV-JvNAhWMOxQKHT1_DK0QIAguMAE&url=http%3A%2F%2Fwebcache.googleusercontent.com%2Fsearch%3Fq%3Dcache%3Amv0AK0akK5YJ%3Awww.mdpi.com%2F1424-8220%2F14%2F2%2F2135%2Fpdf%2B%26cd%3D2%26hl%3Den%26ct%3Dclnk%26gl%3Dus&usg=AFQjCNE-F0XQoTEIH6uJqVZDDzpv_GEt2w&sig2=qvanmJhFDRfaNShjHt7tXw

Hoping this will be helpful,

Rafik

Thank you sir for responce.

Actually what I am trying to do is, I want to immobilized HRP and ABTS in hygroscopic polymer (like Cellulose, Agarose, Chitosan, PVF and so on). I did the test on Agarose but I did not succeed in getting any satisfactory results. What should I do?

And if I am not getting any satisfactory result, what can be the possible cause?

Dear Deepak,

It has been shown that fenugreek hydrogel-agarose matrix with gold nanoparticles is a better film than any of those you mentioned (agarose and etc.). Immobilization of acetylcholinesterase on the membrane resulted in high enzyme retention efficiency (92%) and a significantly prolonged shelf life of the enzyme (half-life, 55 days).

I suspect that your experiments may be were unsuccessful because of the instability of the immobilized enzymes on the agarose matrix.

I suggest that you use enugreek hydrogel-agarose film which consists of  2% fenugreek hydrogel and 2% agarose.

For more on fenugreek hydrogel-agarose matrix with gold nanoparticles, please use the following links:

http://www.ncbi.nlm.nih.gov/pubmed/26320646

Hoping this will be helpful,

Rafik

thank you sir.