Actually i m working on soil urease and stuck at the step of identification of urease activity after ammunium sulphate saturation .i am using sodium hypochlorite rex but i dont know how graph is ploted after taking absorbance.which thing i plot on x-axis and which one on y-axis
Hi. I am looking for this assay too.
If you get one, please could you share it with me? I will do the same.
Firstly, you prepared standard curve for urease by using commercial product of enzymatic reaction and plot different conc. of product on X-axis and their absorbance on Y-axis. After that, add trendline to get the regression and y value for this graph. when u got the y value (y= mx+c), use the value with x as slope of standard to calculate the enzyme activity.
Sorry,
I don't work with soil urease and I don't know anyone assay.
Thanx but can u elaborate abt standard curve more m will be the slope ,,,what equation i use,,,,,,[s]/t=? Plzz reply me soon my work is delaying i will b very grateful to u
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use this equation:
Enzyme activity (U/mL)=Abs. of sample/slope of standard x 1000/Aliquotes (ul) x 1/incubation time x dilution factor
when you prepared standard curve, product conc. should be in umoles. if you prepared in mmoles/umoles solution of product for it, then the value obtained from above equation should be divided with the molecular weight of product used for standard curve preparation. finally you will get U/mL. one unit is defined as the release of 1 umoles of product/minute.
I do not work with soil urease and I work with other enzymes and proteins
one important thing, if u want to show your enzyme activity in terms of soil, then total activity should be divided with amount of soil used for enzyme extraction. then u expressed your activity like U/gm of soil.
Ok thanx,,can u plzz tell me some protocols abt determination of enzymatic activity with some basis,,,actually my supervisior said i need no to draw standard curve rather i should draw o graph of subtrate conc vs velocity(absorption) if i do so than can i use that relation which u told me or for that i search some other formula for the determination of urease activity
it's a same thing. in this condition you will showed the enzyme activity in terms of substrate degradation. if you plot varied conc. of substrate(mg/ug) Vs absorption, absorbance will be decreased in presence of enzyme activity. that equation is common for enzyme activity calculation. Absorbance can be calculted by substracting the abs. of sample with control. Now you will express enzyme activity in terms of mg/ug substrate degraded/minute. you can also show the activity mmoles/umoles degraded substrate by dividing the value obtained through equation with molecular weight of substrate. if required.
Yes so nice of u but i will again ask anything whenever i find any difficulties if u dont mind
It's already known, dilute the desired enzyme solution to get the linearity curve of enzyme at varied concentration.
Do u know any protocol how can we detect zinc in blood sample by atomic absorption spectrophotometry
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