Inhibitory Actions of Pesticides on Alpha-Amylase
Posted: Mon Feb 16, 2009 8:43 pm
Hi,
My project is comparing different rates of inhibition of pesticides on the digestive enzyme alpha-amylase. I have been testing 7 different insecticides, 3 of which are organically based, and the other 4 are synthesized. I have been visually testing this by using a starch indicator I2/KI, a potassium and iodine solution, and I have been timing the reaction of the enzyme+substrate solution till it becomes clear. I am also intrigued about other digestive enzymes such as lipase, and either trypsin or pepsin; however I do not know of a method to visually test these enzymes if you know of a way to do this testing then it would be very useful.
My second question is how I can test this with a spectrophotometer. I have read many papers on Michaelis Menten kinetics however I am still having some trouble understanding how to actually graph velocity of the reaction. Also how should I go testing the reaction of the enzyme+substrate in the spectrophotometer, I am currently using a spectronic-20 and I am having some trouble measuring the absorbency over time and I think a step by step instruction on how to actually perform the experiment in the spectrophotometer would be very helpful. Please tell me if I have left any specific details out of this very short explanation, thank you very much though!
My project is comparing different rates of inhibition of pesticides on the digestive enzyme alpha-amylase. I have been testing 7 different insecticides, 3 of which are organically based, and the other 4 are synthesized. I have been visually testing this by using a starch indicator I2/KI, a potassium and iodine solution, and I have been timing the reaction of the enzyme+substrate solution till it becomes clear. I am also intrigued about other digestive enzymes such as lipase, and either trypsin or pepsin; however I do not know of a method to visually test these enzymes if you know of a way to do this testing then it would be very useful.
My second question is how I can test this with a spectrophotometer. I have read many papers on Michaelis Menten kinetics however I am still having some trouble understanding how to actually graph velocity of the reaction. Also how should I go testing the reaction of the enzyme+substrate in the spectrophotometer, I am currently using a spectronic-20 and I am having some trouble measuring the absorbency over time and I think a step by step instruction on how to actually perform the experiment in the spectrophotometer would be very helpful. Please tell me if I have left any specific details out of this very short explanation, thank you very much though!