i prepared my reagent as follows 30 g potassium sodium tartrate, 1.6 g NaOH, and 1 g 3,5-Dinitro-2-hydroxybenzoic acid. I filled it up to 100ml.
I prepared my glucose standard solution as follows:
0.01 g (glucose powder) + 1 ml, 0.02 g + 1 ml, 0.03 g + 1 ml, 0.04 g + 1 ml, and 0.05 g + 1 ml.
after this i added 1 ml of the DNS solution to each tube and then placed the tubes at 100 C. However, the lowest concentrations appeared to be darker. The standard curve looks backward. I don't understand as in to why the results appear like this and I am really looking forward to your expert opinions.
Well my absorbance value for the lowest tube (0.01 + 1 ml) was about 3.397 and the of the highest concentration (0.05 g + 1 ml) was 0.975. However, I really don't understand as in to why the results ended like this. It should be that the lower concentrated should be lighter and the higher should be darker.
Agreed with Greg. Depending on your spec, you may accept an absorbance as high as 1.5, but surely no higher than that. You're most likely well over your linear range. Good luck.
Thank you guys for your help. I reduced my concentration to 0-1 mg/ml and I can clearly see a difference. However, I am curious as in to why at a concentration such as 0-50 g/L or 0-25 mg/ml the coloring looked exactly the same and with 10 g/L having a darker color followed by 20 g/L because i believe this method should also be able to work at these concentrations. Was there something off in my protocol? by the way after heating i added 7 ml ddH2O.
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