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Bio: Lab: Inhibition effect on Enzymatic Activity

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heya,

i just did a lab on how changes in concentration of inhibitor (lead acetate) affects the activity of catechol oxidase. But i got some strange absorbance readings. The website where i got the idea from said to get the transmission but my teacher said to do absorbance for some reason and i have absolutely no idea what sort of readings i should get, what i got doesnt really match my hypothesis, so i was wondering what sort of colour changes that other ppl got? or if you know a website that sort of tells you what sort a graph you should get?? thanks :P

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Well if you did light absorbance instead of transmission, what did you measure it with? Transmission of light can be measured by comparing the light intensity before it passed through the substance with the intensity of the light after it passed through (lit. how much light was transmitted through something). If nothing was absorbed, you had 100% transmission, if 50% were absorbed then you have 50% transmission and so on. Look it all up for a better explanation, but you're still on the right track, you just have to work out what the relationship is between transmission and absorption and then once you've got that you should be okay.

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Well if you did light absorbance instead of transmission, what did you measure it with? Transmission of light can be measured by comparing the light intensity before it passed through the substance with the intensity of the light after it passed through (lit. how much light was transmitted through something). If nothing was absorbed, you had 100% transmission, if 50% were absorbed then you have 50% transmission and so on. Look it all up for a better explanation, but you're still on the right track, you just have to work out what the relationship is between transmission and absorption and then once you've got that you should be okay.

thanks :S i did that, used this formula to calculate the transmission (or transmittence as its called apparently) But i still have no clue what to make of my readings...i'm getting sort of a...bell shaped graph when i think it should be like small negative exponential ish like curve...i'm thinking of just changing the data slightly?? cos it should the absorption should decrease with higher concentrations of lead acetate (and transmission should increase) right???

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Hrm yeah I'm pretty sure there's no bell-shaped curve in there if all you're doing is changing concentration.

Did you use a light intensity meter to measure the absorbance? Those things're pretty rubbish and it could be that, unless the temperature of your classroom was conveniently flipping along the enzymatic scale as you did the experiment :S You could try making up results, I guess, or if the whole thing is weird it might be a good idea to just repeat it again and see what happens. For IAs you should be doing at least 3 repeats anyway (I dunno if you did any) so maybe doing it again would clear it up.

Also checking your method quickly to make sure you're not accidentally reusing or mixing in substrate, or changing temperature or anything like that. People often forget and start re-using the same equipment without washing it properly, not making sure the conditions of the substrate are fair re:surface area, place of measurement etc., or combining previous solutions with new ones to save time and whatnot.

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lol yes i did repeat them, but then i kind of didnt make new solutions :S i just poured the stuff back into the tubes and poured it back into the cuvettes XD lol i guess laziness is not the way to go :S

and yes i'm making up the readings, and since all the labs are due in on monday, ill see if the teacher can make do with just a design :P

thanks for your help though :S

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lol yes i did repeat them, but then i kind of didnt make new solutions i just poured the stuff back into the tubes and poured it back into the cuvettes :S lol i guess laziness is not the way to go

and yes i'm making up the readings, and since all the labs are due in on monday, ill see if the teacher can make do with just a design XD

thanks for your help though :S

Well that sounds like it explains why it didn't work, if you mean you were re-using substrate! Not good xP

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Hi, I just did this lab too.

Oh my gosh i'm so confused! We measured the percent transmission of the light through the tubes using a colorimeter...

Someone in my class talked about the absorbence of the brown color. What is this supposed to mean? Does it mean that the brown color absorbs the light so when we have a lot of inhibitor and no melanin is formed, that no absorbance occurs so all the light is transmitted?

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